Omics! Omics!

Does Illlmina Also Have A Homopolymer Problem?

2012-01-30T23:38:00.001-05:00

One of the most widely-publicized error modes with Ion Torrent and 454 sequencing has been the challenge of correctly counting the number of bases in homopolymer runs. Because these chemistries use non-terminating nucleotides, polymerase is free to add as many as possible. Unfortunately, the signal linearity breaks down, making it difficult to correctly count. Ion Torrent today released a note on homopolymers, but rather than plowing this well-trod ground it goes for a less publicized problem: Illumina having a more specific challenge in this department. The note is available on the Ion Community, free registration required.

Roche Guns For Illumina

2012-01-25T23:16:00.000-05:00

Due to a business dinner & general exhaustion, I turned in early last night & was caught unaware this morning of the big news: Roche is making a hostile takeover bid for Illumina. Ugh!
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Sequencing Technology Fireworks

2012-01-10T22:52:00.000-05:00

I actually awoke today expecting an exciting press release, but I sure wasn't prepared for the big announcements from Ion and Illumina. Not that they were totally unexpected, but there's a huge difference between speculation and announced products (which, of course, are hugely different from ones you can actually buy!)

Ion Torrent Pairs: To What End?

2012-01-07T23:13:00.000-05:00

Ion Torrent quietly released a set of paired end datasets over the holiday break. This is a bit embarassing for me, as in my last post on Ion I stated the platform "will probably never have paired ends" and in fact Ion had already announced the protocol. Oy! I also missed their mate pair protocol being released, though the document itself is another victim of Ion's incredibly counterproductive security policy. If you don't own a PGM, you can't access the document -- never mind if you are trying to plan for a potential buy or are preparing a library for a friend/collaborator to run.

Year's End

2011-12-28T13:10:00.000-05:00

I hoped this year to push myself to blog more frequently and regularly. Clearly I did better than some years, but not up to the standard I had hoped for. I've also realized that I missed noting some significant personal milestones.

Reflecting on a Year of Ion Torrent

2011-12-09T08:46:00.001-05:00

Ion Torrent released three more datasets this morning, all generated on their 318 chip. One's from E.coli but two are human genomic samples. With approximately 1.2Gbp of raw data coming from these 318 chips (fron around 6 million quality filtered reads per chip), they are starting to move up the food chain in human genomics from pure amplicon sequencing to more complex small targeted resequencing efforts.

Fitting Out

2011-10-27T08:37:00.000-04:00

One of the attractions of my new shop was the possibility to see a biotech company built from the ground up. Each of my previous companies had been a long-standing concern by the time I got there; even Codon Devices had a year plus under its belt and some equipment already mothballed. The new venture moved into its first lab space last week, and as you can see from the picture all we have at the moment there are bare walls.

MiSeq Made Easy?

2011-10-11T06:01:00.001-04:00

The first computer I ever tried to program was built from a kit by my brother and father. The DATAC-1000 was a single-board machine, with that single printed circuit board about the area of a large laptop (image on page 9). Sporting a grand 1K of RAM, it was a grand machine. User input-output was entirely through a set of binary touchpads and LEDs, though a cassette tape interface enabled storing and reading programs. If I helped any with it, I might have sorted the resistors since I had just learned the color code. The machine sported the same processor as some other machines of the time, such as the KIM-1 and the PET and even something called an Apple.
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Thinking Outside the Box or Just Plain Nuts?

2011-09-28T23:52:00.000-04:00

Please take the title in the spirit it is intended: as a bit lighthearted. Seeing the object pictured and reading the accompanying blog post from one of Jonathan Eisen's graduate students. It's an unusual solution to a common problem, and gave me a good chuckle.

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Boston's Boris Bikes

2011-09-27T21:52:00.000-04:00

When I discovered that my new gig would temporarily in Boston, I realized I had an opportunity to try out Boston's new bikeshare program. Started this summer, Hubway consists of racks of bikes in public places which can be used for short hops around town. I like my folding bike, but on some rush hour trains it is very hard to find space for it, especially with some train conductors who are more interested in giving dirty looks than serving their passengers. Plus, it's now quite dark on the last leg of my commute, and even if I had some really slick lights I don't like riding even short distances in the dark.

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Transitions

2011-09-18T21:55:00.000-04:00

I went to an Infinity going-away lunch last week. We head off to some favorite local restaurant and order a modest (but delicious) meal on the company dime. The departee makes an impromptu speech, there are goodbyes and handshakes and usually a number of pleas to stay, both fictitious and heartfelt. Those staying wonder what could lure someone away from the very safe and green pastures of the company. I've been to many such lunches with Millennium and Infinity; with Codon the lunches tended to be group affairs as people were laid off in batches.

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Illumina Calls for a Flag on the Play

2011-09-14T09:10:00.001-04:00

Continuing my sports analogies, but switching games, in my coverage of the benchtop sequencer war today. Alas, I can't refer to instant replay, as the usual set of procrastination excuses has resulted in this being filed very late after I was made aware of it (first by a comment in the blog, then by a friendly chap from Illumina alerting me). In any case, Illumina has responded to Ion Torrent's claims on long reads and overestimated MiSeq quality, and mostly done so by crying "Foul!".
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Genetic Education:

2011-09-01T08:30:00.093-04:00

A study by the American Society of Human Genetics (ASHG) is published today suggesting that few U.S. states have adequate standards in place for genetics education. I forgot to ask for a link to the article that would go live post-embargo, but it should be on the website of the journal (CBE Life Sciences Education)

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Will Max-Seq Gain Traction?

2011-08-31T22:59:00.000-04:00

At the beginning of the month, I had dinner with J Adams from Azco Biotech and some friends/colleagues of his and talked over the Max-Seq. And yes, I did let them pick up the bill -- J wouldn't let me pay for myself. In Sequence did a nice piece on it the next week, so again I've blown an opportunity to scoop them. Then somehow, between vacations and other distractions, this piece was stuck in blogger limbo. But, there are some details I don't see there and some color I think worth adding.

Wishing I Had Been A Referee: A Renaissance for Tagamet?

2011-08-28T08:42:00.000-04:00

Back from vacation & watching another wave of Hurricane Irene soak the area (curiously, the windiest times so far seem to be breaks in the rain). August has not seen much attention paid to this space (indeed, I have one piece that has gestated nearly the whole month), so time to put the shoulder to the wheel.

Just before my vacation, a pair of papers showed up in Science Translational Medicine which describe two attempts at drug re-positioning by transcriptional profiling. The key concept is to take expression profiles for diseases and try to find drugs which appear to generate the opposite transcriptional pattern, with the theory that the drug could nudge the disease pattern back to a normal state. This is an idea which has been kicking around for a while, and at one time was the focus of a number of companies. One was even trying to re-position a drug I have a small connection to (MLNM developed it from a target I spotted in an EST library), but I believe that is a dead effort. One challenge in tracking this field is that it is rarely obvious what happened in the end; did a drug fail to pan out or did the backers just run out of cash?

Names in Collision

2011-08-08T22:59:00.000-04:00

I will claim that I saw this coming, in that I've toyed with the basic skeleton of this post before. But, I hadn't gotten around to it -- but how could I miss this opportunity. On a mailing list devoted to SAM, someone asked about a related topic to SAM, and one of the experts on the board replied with an electronic head-scratching

Ion Throws A Long Punch At MiSeq

2011-08-04T23:17:00.003-04:00

The benchtop sequencer wars are heating up! Illumina and Life are engaged in a fierce war of pamphlets and datasets to convince the world that they have the edge. I won't attempt to give a complete play-by-play, but hit on the latest developments, which includes Ion releasing a dataset of 250+ bp reads.
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How Many Toes Does Ion Have Left?

2011-07-29T09:21:00.000-04:00

I've previously complained about Ion Torrent's bungling and secrecy when it comes to educating their current and potential future users about key technical information. I've recently come across yet another botch, one that underscores that attempting to control information in the Internet age only serves to distract from the goal of ensuring access to correct information.

Tschuess, Shuttle!

2011-07-28T08:55:00.000-04:00

Clearly with a week having passed since touchdown, my farewell to the space shuttle program is grossly overdue. I actually missed watching the event live, as I was off on a camping trip in some beautiful New Hampshire woods.

Atop of the usual excuses, it's been a bit hard to figure out what I could write that was different than so much that has been written. It would seem that every major media outlet opined on the subject, and most did a decent job in hitting the key points.

Ion's Gaggle of New Accessory Products

2011-07-12T00:11:00.000-04:00

Ion Torrent had a press release announcing a number of new components to the platform, as well as shipping of the 316 chip and a drop in price of the original 314 chip to $99 (alas, the announcement was not synchronized with updating the online store, so no details could be checked -- such as the price of the 316 chips).
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PacBio's Foray Against the German E.coli Outbreak

2011-07-11T08:53:00.000-04:00

I recently remarked that it was surprising that PacBio had not jumped on the German E.coli outbreak strain, given CEO Eric Schadt's professed interest in biosurveillance. Dr. Schadt was kind enough to spend a half hour discussing the topic with me last week, in the wake of PacBio releasing de novo assemblies of this strain and 11 other pathogenic E.coli strains (6 having never previously been sequenced) on the PacBio DevNet website. Dr. Schadt also has a detailed blog post on the project.

In both the blog post and our conversation yesterday he explained why PacBio wasn't in the forefront of this effort but still decided to jump in. Dr. Schadt was certainly aware of the outbreak; he was in Germany at a conference when the public alarm was building. But initially, the company was trying to stay focused on the commercial launch of their instrument. However, after seeing the first public assembly come out at several thousand contigs, they consulted some academic collaborators and decided to run their own sequencing, with a goal of providing a much less fragmented assembly. The initial thought was a hybrid assembly containing both PacBio and short read data, but after another group generated a high quality short read assembly the emphasis switched to a PacBio-only assembly.

Sequencing proceeded rapidly, as before; about 2 days from receipt of the samples to generation of data for assembly. Three machines were enlisted, using current generation SMRTcells but with pre-release polymerase and protocols (these are scheduled to go to customers by year's end). A number of eye-popping statistics are shown: mean mappable reads of 2.5-3Kb, 5% of the reads at 6-7Kb or greater and one herculean read of almost 23Kb! Yield from the SMRTcells was variable, ranging from a mean per sample of 13K to 55K (overall mean of 26K). Raw accuracy was still about 85%.

For the German strain, both a large insert library and a small insert circular consensus library were sequenced. One challenge that the PacBio team ran into is rapidly generating a DNA population of defined size which is bigger than about 9Kb. The populations must have a restricted size range, or else the smaller fragments are preferentially loaded into the zero mode waveguides where the sequencing actually takes place. Due to this, the value of the monster reads was diminished. Indeed, Schadt sees this leading to PacBio's inability to drive their data to a single contig (the main chromosome is broken into 33 contigs); there simply weren't reads able to bridge some very large repeat elements. This also is why they didn't use the strobe sequencing mode, which generates islands of sequence separated by statistically-defined gaps. Given that so many of the reads were approaching the size of the inserts, strobing wouldn't do much good.

Not very far back, I presented a somewhat bearish case for PacBio. Do the latest results change this? As much as I love the technology and the idea of long reads, I'm still concerned that too many scientists will see these as a nice-to-have and not a must have. Dr. Schadt mentioned they are working on projects to demonstrate PacBio sequencing of much larger (100+Mb) genomes, which is an important start. Still, it may be that few labs will see the incremental value of long PacBio reads as not important enough, or that they simply don't need to invest in a machine but instead rent some of the existing capacity (I know of two service providers offering the system). As PacBio pushes the reads longer and longer (imagine if 5% of the reads were 20+Kb!) it will offer advantages for closing long gaps. For example, PacBio should require relatively little DNA for library construction, whereas some of the competing mate-pair techniques are notorious for being very inefficient at converting input DNA to usable fragments (as well as creating some level of noise from chimeras generated in ligation steps).

MiSeq Aims for Ion Torrent

2011-06-24T15:15:00.000-04:00

Illumina has released a MiSeq dataset for E.coli MG1655 on its website. Accompanying the FASTQ and BAM files is a presentation, the first half of which compares the performance to big brother HiSeq. The second half is an explicit comparison against the available Ion Torrent.

A first peek at data from an Ion 316 chip

2011-06-21T23:33:00.000-04:00

Ion Torrent has made available an E.coli DH10B fragment dataset for the 316 chip, which is expected to be generally available early next month. I've theoretically had access to the data since Saturday, but a series of events (computer-free weekend, a crashed home computer, a personal day & a good but unbloggable conference) have meant that ambitious plans to analyze it are still in progress. So, what I discuss below is either from Ion's provided information or other sources, so take it with appropriate caution.

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Two administrative notes

2011-06-21T23:31:00.000-04:00

Two administrative items:

First, sometimes comments are getting moderated very late. I do pledge this is not an attempt on my part to bury negative commentary. Rather, it appears that sometimes when I thought I had moderated comments via my Android phone it hasn't succeeded.

Second, I am trying out the new smartphone-friendly layout option for Blogger. Please let me know if you are reading this on a smartphone and hate the new layout.

E.coli Outbreak Genomics

2011-06-15T23:24:00.001-04:00

The E.coli outbreak in Germany continues to be a major news item. It is looking increasingly doubtful that the source of the infection will be conclusively traced, as the German authorities have already named and then backed off two suspects, Spanish cucumbers and German bean sprouts. These activities have not been without repercussions; Spanish agriculture has been hard hit and exports of European produce in general are reportedly hurting.

On the genomics front, the outbreak has demonstrated how quickly bacterial genomics can be run on the current class of instrumentation. BGI Europe knocked off the sequence in a series of Ion Torrent runs in 3 days, and a group at University of Muenster worked at similar speed with the Ion setup as well. Later, sequences have come in from the Illumina and 454 platforms. The public release of this data has engendered a number of public analysis projects.

The Arsenic Bug, Revisited

2011-06-05T22:01:00.000-04:00

As has been covered in many outlets, Science has released eight technical comments on the arsenic-loving bacterium issue along with a response by the authors, which I commented on when it appeared. A good summary of the comments can be found at In The Pipeline, and Fejes.ca has a good take on it as well. The quick summary is that various technical and theoretical issues are presented by the critics, and the NASA team dismisses all of them by standing by their data and interpretations.

Paying a Painful 75% Secrecy Tax

2011-05-26T23:57:00.000-04:00

In a post a while back, I mentioned that my Ion Torrent sequencing project was stalled because my service provider couldn't get some of the key kits, despite an Ion representative posting that no such shortages existed. I've been remiss in updating that; last Tuesday the kits showed up and Monday I got my data -- and a bit of a shock.

MiSeq's First Light

2011-05-23T22:31:00.000-04:00

Someone was kind enough to send me a copy of a poster by Illumina reporting results from the MiSeq. Now, to be very upfront, by someone I mean "a person from a PR firm contracted by Illumina" and "kind enough" that she was doing her job. I don't have illusions about motive here, the author list is all from Illumina or Epicentre, but this was a poster presented at the recent Cold Spring Harbor Biology of Genomes meeting. It certainly isn't peer-reviewed data, but it is something. Of course, we can't know to what degree these are cherry-picked results. If you want to be really cynical, call it messaging and not data. Yes, I've taken some flak in the comments recently about how favorable my coverage of Ion has been, and I'm trying to adjust. And don't worry; I have some new bones to pick in that area.
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Forums: Open Beats Closed Hands Down

2011-05-19T22:22:00.000-04:00

Around the Internet, there are a number of communities in which scientists can swap useful information. SEQAnswers is a very useful site I frequent; BioStar is one I don't but probably should. Life Technologies has set up a community around Ion Torrent, and the contrast between that and SEQAnswers is a useful one.

SEQAnswers has a straightforward access policy. Anyone can view content, but to post or reply you must register for a membership. This approach appears to have been very successful, as there is a healthy number of individuals posting to the site. You can browse around and figure out if the site applies, plus search engines such as Google can steer individuals in. SEQAnswers boasts a number of authors of major second generation sequencing analysis packages, including Bowtie, Tophat, BFAST and Samtools, as regular contributors. There is a significant network benefit to this; quality encourages quality and conversely such folks must be judicious in the number of forums they actively participate in. The management of SEQAnswers applies a light hand, occasionally moving posts to more relevant forums and smothering all spam. In addition to the forums, a key asset is a large wiki on second generation sequencing packages.

The Ion Community is set up on a very different basis. It has two sections, each with its own membership restrictions. PGM Users is open only to registered owners of the sequencing system; Torrent Dev is open to that group plus anyone registered for the Grand Challenges. Each section has both discussion areas and documents. The site is flashy, though more than a few links are indirect detours to what you really want.

Now, there are plenty of examples of how maintaining some control over a site can be productive. I was recently trying to eradicate one of these nefarious fake antivirus viruses from our home computer, and on one major security software company's forums I found what looked suspiciously like a link to infect with one of these viruses. Keeping a single point of origin for documents can be useful as well, to reduce confusion. For example, if you Google around for information on 454 fusion amplicon design it is easy to find outdated information. We also wouldn't want any forum to devolve to the level of the Biotech Rumor Mill, which by its very nature must allowed unregistered posters, and as a result is a mudpit of insults and near(?)-libel.

But, Ion's approach is in my opinion strongly self-defeating. I won't go into detail here, but in the extreme form they have on two occasions (this thread and this one) argued for the suppression of information on PGM from SEQAnswers (I will try to tackle this soon, but after getting a chance to talk to at least one person with the Ion side of the story). But it's also easy to argue from a purely practical standpoint, rather than a philosophical one, that this approach is not doing their platform any favors.

The first problem is that the closed access means that you can't lurk on the cheap there; either commit to being part of the community or stay out. This prevents sucking people in slowly; for many the barrier of registering -- particularly since your registration does not become instantly active -- is too high a barrier. Indeed, I would offer as evidence for this that the first major PGM-tuned software package not from Ion or a commercial partner was apparently not spurred by the Ion Community, but by Nick Loman's post on assembly of Ion data. Nick's original post apparently received over 2K views, which must be at least an order of magnitude larger than the current Ion Community membership.

The second problem is the two layer design. Okay, I'll admit it -- it's maddening to be excluded from the PGM Users forum. How do I know that there are not discussions there I could either benefit from or contribute to? If someone starts a discussion there better suited for Torrent Dev, will it get bumped up? If so, who decides? But, worse than that is that many technical documents that I would find valuable are beyond that access barrier.

The third problem is I find these rigid definitions completely at odds with scientific reality. Just because I don't own a PGM doesn't mean I might try to do everything but run the instrument. As an example on another platform, my colleagues & I have run SureSelect on Illumina where we did all the steps except shear the DNA (outsourced), prep the flowcell and run the flowcell. Furthermore, in a modern collaborative environment, someone in one lab may own the machine but in another lab work up the samples. It's also not clear what any of this "security by obscurity" is buying; given the number of groups using Ion, whomever they're trying to hide the information from can certainly find a leak. Ion should heed the example of the music industry, which failed to provide legitimate means to supply a growing demand for digital music, and thereby spawned widespread illegal file sharing. Plus, many browsers are potential buyers -- people want to know what they are really buying in to and to start storyboarding what running an instrument would mean in terms of personnel and auxiliary equipment.

The fourth problem is discoverability. How do I find information? Well, for second generation sequencing stuff it is the trio of PubMed, Google and the SEQAnswers software wiki. PubMed is great, but many packages show up online long before they are published. Google is useful if you know what you are looking for, and SEQAnswers is great if someone has logged it there (and in general, once I see something I make sure it is logged there).

But with the Ion Community, those last two are problematic. The objections Ion has raised to links going to the interior of the community mean that I don't dare put such in the Wiki; but conversely a link just pointing to the community is not terribly useful. But worst, since Ion apparently won't let Google in their stuff is invisible to that valuable tool. As evidence, at the moment if you Google for information on their TMAP aligner with "tmap source code ion torrent", nothing from the community comes up (but threads on SEQAnswers do!).

Ion isn't the first, and probably won't be the last, company to try to have its I proprietary control cake yet eat its Internet openness. Life has SOLiD Community, Helicos has one for Helioscope, PacBio DevNet for SMRT sequencing and so forth. It takes some real courage to relax some control and invite the whole world to your party. Some folks would no doubt see view-without-registration as a loss of useful marketing data, forgetting that the openness of a site will itself draw in customers. Anyone launching a new platform should really ask themselves, will I be better off trying to control a rare destination for a few visitors, or perhaps just cultivate a sub-community over at SEQAnswers.

Oh Would It Be Fun to Debug Strobe Sequencing!

2011-05-14T22:30:00.000-04:00

Through the course of a month, I easily sketch out (on average) a dozen plus experimental designs in the context of my day job. Of course, only a very few of these are ever executed; many never get shown to another soul. By constantly pondering how I might tackle questions, I can keep in practice and present plans which are reasonably well thought-out. It also helps to have thought through things; sometimes a rejected plan will suddenly look like a gem due to some other result or change in priorities.

Atop that, I also sketch up a few which have nothing to do with my day job. Partly it is fun, and partly it is a way to exercise the process on areas I can be certain of my objectivity. A related exercise is to sketch out a business plan; I end up doing that a few times a year. Never have taken any further than a napkin; not only do I lack the necessary thirst for risk, but most are for businesses I'd be happier being a customer than an employee.

For example, after a recent lunch with a friend from graduate school I found myself again contemplating a question that had arisen at Codon in the context of gene synthesis. We didn't have second generation sequencing there (if Codon had survived, we certainly would have launched into it at some point) and had seen the hint of an interesting phenomenon (and practical problem) but could have never collected enough data to really nail it down. Now, with sequencing cheap on a large scale, and this being the perfect sort of problem for such sequencing (since one would need only very short reads), it was short-term obsession to work out how to run such an experiment. For probably $5K and a tiny bit of molecular biology, a nice little paper -- pity I don't have a slush fund to cover it. My apologies for not supplying any details; maybe I will someday have some found money to cover the project.

However, I did just get another idea that I might as welll be open about, as for one it will probably be solved in the near future and two there is no opportunity to actually work on it. So it can be fun to speculate in the open on how to address the problem.

Blogger glitch

2011-05-13T16:47:00.000-04:00

Blogger experienced an outage which I found out about today. This caused my last post to disappear; any posts or comments in a certain time period were lost.

Thanks go out to the anonymous reader who posted a comment wondering where my last post went. Blogger (Google) did not notify me of the issue. The really disconcerting part is the Google cached version of the post was missing as well -- this suggested something very odd going on.

I often write posts directly in Blogger, so I don't have a backup. This was an exception & I did actually have the rough draft, which I would have posted if the old one wasn't restored.

Ion's Growing Pains

2011-05-11T23:44:00.000-04:00

A recent In Sequence article indicated that Ion Torrent is enjoying strong initial sales. This bodes well for continued evolution and improvement of the technology, as LIFE will continue to smell revenues and opportunity. Ion has announced a number of improvements, but most aren't scheduled to arrive until the near future.

The challenge is for LIFE to keep executing on their plan. Already some issues have arisen; my own Ion experiment at a service provider is stalled due to a back-ordered template prep reagent (two weeks and counting!). This is a key reason to do pilots on emerging technologies with non-critical (but interesting) samples; I was bitten last fall by another backorder bug (that time, the paired end SOLiD reagents). Of course, this time it is even more complicated, as Ion is making a major change to both the underlying kits and to the software to process the data. It will be worth it if I get results anything like Ion's provided E.coli 314 dataset, which has about 4.5X the data of the original 314 chip spec.

Ion Torrent's Data Quality Is Pretty Good (and Better Than Ion Claims)

2011-05-08T22:12:00.001-04:00

One of the key questions around Ion Torrent, as with all new platforms, is what is the sequence data quality like. Now, that can be a loaded question but I'll ask a slight variant on it: how truthful (or accurate) is Ion at estimating base quality?

Quality scores are a useful adjunct to sequencing data and are commonly expressed as phred scores, which are the integer part of -10*log10 of the error probability. Any base caller needs to estimate these and many downstream programs, from aligners to assemblers to variant callers, rely on these quality values for their operations. In many cases, the individual quality scores are combined to generate some joint estimate of the error (I built one such model at Codon). These error probabilities come not from an infallible source, but are rather estimated from aspects of the raw data.

Which numbers did I use this time?

2011-05-07T23:01:00.000-04:00

Noted screenwriter William Goldman's second memoir on the film industry is titled "Which Lie Did I Tell?". The title is not a quote from Goldman, but rather what another movie industry said after getting off a long phone call he had taken in Goldman's presence. I'm a bit nervous I inadvertently strayed in that direction in my last item on Pacific Biosciences.

I was relying on memory for my numbers & in the course of writing things I think I also revised those downwards, not out of malice but rather an attempt to be conservative. As a correspondent pointed out, the first pass accuracy on the first commercial system is claimed to be 85%, not 80% as I stated. On the number of reads, I failed to update for the newer SMRT cells which are out; I said 10K and it's probably at least 3X that. However, I do have a bone to pick.

How Many More Machines will PacBio Sell This Year?

2011-05-02T20:37:00.000-04:00

Amongst last week's news is the item that Pacific Biosciences has officially launched their SMRT sequencing platform. I'd too eye-deep in various projects to figure out how off schedule that is (I think nearly a year from their original target), but now it is launched.

Better Template Prep for Ion Announced

2011-04-13T09:31:00.002-04:00

One of the perceived weak points of the Ion Torrent, in contrast to the Illumina, has been the use of emulsion PCR for template preparation. The original template prep protocol was apparently around 10 hours of wall clock time, with a substantial amount of hands on time. Improved template prep is the subject of one of the Grand Challenges. A new kit being released today along with a new instrument announced today (but not generally available until late summer or sometime this fall) go after this issue; the new kit providing an inexpensive but substantial immediate improvement and the modestly priced template prep instrument providing a very low labor solution once it arrives

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Vostok & Columbia

2011-04-12T09:05:00.000-04:00

It has hardly gone unreported in the media that today marks the 50th anniversary of the manned spaceflight and the 30th of the first space shuttle launch. It was an accident of scheduling delays that put that flight on the 20th anniversary of Gagarin's, but what an appropriate synchrony that is! My own contribution herein is to pen two quick capsules of two books that deserve longer reviews: Two Sides of the Moon and Riding Rockets. If you have a strong interest in the history of spaceflight, you should consider reading each of these if you haven't already. The only caveat I'll throw in is that if you have a young friend who fits that category, but you do not, you should at least skim Reading Rockets before passing it on; some of the content (and most of the humor) is of a mature nature. While neither book is primarily about either of the events commemorated today, both bear on it.

Gnoteworthy But Gnot Gnoticed

2011-04-08T22:19:00.000-04:00

A commenter on yesterday's piece on Intelligent Biosystems scolded me on not mentioning GnuBio and said they had released data. This had totally slipped my my notice, and indeed it seemed to have slipped past the GenomeWeb and BioIT worlds as well, judging from some Google searches. There is certainly a press release out there and covered by several outlets, but amazingly stealthy for public relations. Strange! But many thanks to my anonymous correspondent for flagging me on this!

But, it does make a set of stunning claims. When GnuBio launched last year and announced they would have alpha instruments in collaborator's hands by the end of 2010, I was skeptical.
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Another Low Cost Sequencer on the Horizon?

2011-04-08T00:13:00.000-04:00

An article in GenomeWeb's In Sequence (which, alas, requires a subscription for which I've never sprung) has a piece on Intelligent Biosystems (IBS), which iat the X-Gen Congress meeting apparently announced a plan to launch the "Pinpoint Mini" sequencer. The box would come in at $85K, putting it smack in the middle of Ion Torrent PGM and Illumina MiSeq pricing. The hope is to have boxes shipping to early access customers by the end of the year.

To be honest, I'm guilty of mentally writing off IBS, as they had been around a long time and very quiet. Indeed, in my defense their website looks like it hasn't been updated since it first went up, and you'd think now that they made a big announcement it would be updated, but apparently not yet. On the other hand, while stale websites can indicate fading companies, the fact that it still is working suggests some life.

In any case, I was apparently hasty in my thoughts. The box as described has some interesting features. Supposedly it will crank out data at $75/Gbase. The claim is that one exome could be sequenced (reagent costs only, mind you) at 30X for $150 in about 1.5 days. No word on read lengths. The system will mount 20 flow cells, each of which can be run independently. Chemistry is based on reversible terminators that they exclusively licensed; if I understand it correctly the big advantage of their chemistry is simplicity. There's also a curious bit in the publication from the founders is using a mix of unlabeled reversible terminators with labeled dideoxy terminators; the same cleavage reaction removes both the terminator and the label. This was touted as a way to reduce the discrimination of the polymerase against the reversible terminators. Of course, an alternative would be to generate mutant polymerases which are more amenable to being fed terminators. Having four pairs of complicated compounds wouldn't seem to be a route to low cost, but perhaps the gains are worth it (or this chemistry isn't being used any more; very hard to tell from what I was able to read).

Sounds great, but of course there's a lot to do before such a machine can launch. There's no word about sample prep method, which probably means it will use emulsion PCR, since necessary licenses for that can apparently be obtained. There's also the problem of manufacturing the instrument and the reagent kits. Te fact that IBS apparently planned to launch a PinPoint large-scale sequencer a few years back and couldn't get it out the door is not going to help them compete in the expectations market with the other boxes.

One solution to some of these issues would be a strategic partnership with (or outright acquisition by) a major reagents and/or equipment player. It's not hard to come up with a list of candidates, based on nothing more than that description. Perhaps at some point Affymetrix will decide to move into next-gen. Roche could always decide to go for something cheaper than 454, but I doubt it. Agilent seems quite happy supplying picks and shovels, but perhaps they'll go for the big time. Perkin Elmer, GE (which is working on blue sky sequencers) or a host of others. Picking the right partner will be key; Illumina has the advantage of an enormous installed base and thriving ecosystem of associated vendors, whereas Ion Torrent has a lot of buzz and serious marketing muscle (not to say Illumina is lacking there either).

It's also interesting to see this machine being touted as an exome sequencing workhorse for clinical use. The issue really deserves its own detailed post, but such an application brings some serious issues. Library preparation requires a lot of labor and a bunch of other instruments, or a bit less labor and some more instruments specialized for prep. Right now, the market for exomes on HiSeq using either SureSelect or EZCap is quite competitive; I've recently gotten quotes ranging from $2K-$5K (the lower quotes tend to be from new entrants; promised coverage varies a bit too) . For 50Mb capture at 50X coverage, it would seem you could get around 50 exomes into one HiSeq flowcell, which at $10K each means about $200 in sequencing (feel free to correct my math in the comments). That would suggest that very little of the cost of these exome captures is sequencing reagents; the majority is labor and the EZCap or SureSelect kits.

While some elegant library-free methods (really, methods which add the sequencing adaptors as they are capturing the targeted DNA). for exome sequencing have been published, none of these are commercially available on an exome scale. RainDance requires an expensive ($200K+) box and isn't quite up to exome scale,. . Halo Genomics. has announced a library-free prep for "1000s of exons", so perhaps this will break this problem open. Whether this is really whole exome, or something smaller, remains to be seen. A safe rule, though, is that these methods are only efficient if read lengths are significant. At a minimum, the first part of a read is burned on getting through the targeting primers, and with very short reads the size of each targeted amplicon must be small, meaning for a fixed number of amplicons (cost) you can capture a lot less DNA than with a longer read technology.

So, another player in the field -- but with a far off beta release and a lack of a track record. They'll be fun to watch (assuming they go out of possum mode), but probably won't be a real factor in the market for over a year.

Can we treat the kinase du jour?

2011-04-05T00:21:00.000-04:00

For the second time in just over a week, the Boston Globe Sunday was discussing protein kinases in the context of cancer. A group from the Broad has just published a sequencing study (Sanger!) identifying mutations in the protein kinase DDR2 in about 4% of squamous cell carcinomas of the lung. This is a common form of smoking-induced non-small cell lung cancer (NSCLC) and one for which many therapies useful in lung cancer are contraindicated. The prior study by another group at the Broad was published a bit over a week ago in Nature detailing an extensive look at myelomas by sequencing, and found mutations in the kinase BRAF in 4% of myelomas.

The myeloma study is quite a watershed and in some ways raises the bar for cancer genomics publications. Whereas most papers have published a single cancer genome and a few have published single digit genomes, this one looked at 38 myelomas. Now, not to overstate things, as only in 23 patients were matched normal and myeloma whole genomes sequenced; for the other 15 patients just the exomes were sequenced (one additional exome pair was run in a patient with whole genome sequencing, to enable comparison). Clearly this is a serious scaling up of effort, enabled by dropping costs. By sequencing multiple genomes, the possibility exists both to discover rare variants as well as get some rough mutation frequency information.

The myeloms study is curious in one aspect: the results were first discussed about a year ago at AACR, the big pre-clinical meeting going on right now, and were described as submitted at a conference I attended at MIT last June. Indeed, the paper states "Received 11 June 2010; accepted 17 January 2011". While there is a bit of functional investigation of one gene (siRNA vs. HOXA9) and some Western blots of coagulation factors, this is primarily a genomics paper. Is Nature becoming reluctant to publish such papers? What really held this up for so long?

In any case, the primary finding in both of these papers is low but measurable frequency mutation of protein kinases in human cancers. This should come as no surprise, as a previous paper from the same groups in lung adenocarcinoma (the other major class of NSCLC), multiple kinases were found to be mutated beyond the relatively high frequency EGFR, again including BRAF but also a host of other kinases. The new DDR2 paper also found mutations in multiple kinases, though any follow-up was focused on DDR2. Another 5%-or-so slice of adenocarcinoma carries a fusion protein of the kinase ALK, which can be treated with inhibitors developed against ALK. It also may be an opportunity to target ALK by a different strategy, one which my company has explored (yes, I have a financial interest there!).. The challenge is to determine which, if any, of these mutations are driving tumors and which are just passengers.

In the case of the DDR2 paper, the authors built a pretty nice story. One big bonus to protein kinases is that there has been extensive efforts in the last 30 or so years to study them, with many inhibitors available. A raging argument in the field is whether clinically useful inhibitors need to be exquisitely specific or can be as subtle as a wrecking ball, and the truth is that clinically approved kinase inhibitors run the gamut. Imatinib (Gleevec) is quite specific, though it still hits multiple kinases and that has proven useful as it has enabled targeting multiple cancers. For example, some gastrointestinal stromal tumors are driven by c-KIT mutations and others by PDGFR mutations, but luckily imatinib hits both. Other inhibitors such as sorafenib and suntinib are less discriminating, but still tolerable.

In the case of DDR2, the approved inhibitor dasatinib turns out to be effective, and the new paper shows this first in cell lines. Cell lines carrying DDR2 mutations are more sensitive to dasatinib than those which do not, but the trend continues both in mouse xenograft models and finally in a single human patient carrying a DDR2 mutation in her tumor. Alas, the patient apparently had to discontinue therapy due to side effects.

Now that genomics has demonstrated the ability to find these low frequency mutations, the question is quite open as to how to move them into clinical practice. One model would be to simply sequence extensively and treat each patient by the best guess for their mutations; this approach has been published and is apparently being used in the case of author Christopher Hitchens. While whole genome or exome sequencing might be too costly or slow for routine use, targeted mutation panels are another possible approach (though honestly, exome sequencing is getting down in the $2.5K range these days). Such targeted panels can attempt to focus on the most frequent and actionable mutations, though DDR2 in squamous cell carcinoma appears to not have any one mutation particularly favored.

The alternative is to try to run clinical trials to carefully appraise the clinical utility of these approaches. When I mentioned the BRAF in myeloma story a while back to a co-worker (who happens to have developed multiple drugs, including an effective one in myeloma) and expressed the opinion that it is a slam-dunk to use a BRAF inhibitor (which is near approval in melanoma) in such cases, he took a more cautious view. How do you know these are really the important mutations? How will you know how long the treatment lasts? Perhaps the BRAF mutations in myeloma help the tumor but are not critical. How will you know the correct dose schedule? Combination therapy? Whether drug is getting to a very different tumor? To truly answer these questions rigorously, trials are needed.

But the difficulty in running such trials cannot be underestimated. For example, a company thinking of running a clinical trial looking at BRAF in squamous cell carcinoma faces quite a task. Now, the market is not small: according to Wikipedia (an easy lookup late at night, though perhaps with large error bars) there are about 500K new cases yearly, and a quarter of those are squamous. Presumably at least a quarter of those cases are in the U.S., so around 70K new cases per year. Four percent of 70K (error bars growing with each estimate) is 2.8K patients, which is possibly attractive but getting small..

However, to get the trial going you are going to need to screen to find that 4% of patients. Squamous is a standard diagnosis, so you can start there, but will still need to recruit, consent and screen to get that small fraction. In the mean time, you are competing with every other trial out there to recruit, consent and screen patients. Sure, once they miss another trial they might come to yours -- or might not. To top it off, a lot of patients either are never offered or will never consent to a trial; the farther you are from a large academic cancer center, the less likely you will have a trial available to you.

Now, if oncogenic mutation screening becomes a standard part of cancer care, as it has at MGH and probably some other leading institutions, then if these mutations are in the panels it may be that many patients will know their mutation status before you recruit them into your trial. But until this becomes widespread, and only if your gene of interest is sufficiently covered, will this method work.

Yet another approach is to design trials which test multiple therapies. One prominent example in lung cancer is the BATTLE trial, which is trying 4 different therapies with an adaptive design which uses molecular testing as part of the therapy-assignment scheme. Designs such as BATTLE are quite complicated (well beyond my expertise to critique) and get only more so with more drug regimens; if lung cancer is driven by a dozen or so kinases suggesting a slightly smaller number of therapies, can a trial to test these therapies be designed, patients accrued and useful results out? In such studies, will they be judged by whether the study overall improves survival, or can each treatment be viewed as a separate study?

For the sake of patients, these issues need to be tackled. They'll be hard in lung cancer, and far worse in a disease like myeloma. If we ballpark myeloma at 15K new cases per year in the U.S., 4% of that is getting to be a small group (600 patients). Any sizable trial is going to need to recruit a huge fraction of these patients. Now, with patient advocacy groups and publicity it may be possible to find that small population, but it will certainly be challenging. Indeed, the Multiple Myeloma Research Foundation (which sponsored the sequencing) is already talking about how to support such efforts.

So, in closing, these sequencing studies are suggesting very real therapeutic options for patients. However, driving these findings to routine clinical use, even when drugs are available off-the-shelf for the kinases of interest, will continue to challenge all of the scientists working on translational oncology research.

What might a PGM2 look like?

2011-03-23T23:20:00.001-04:00

The New England chapter of the Laboratory Robot Interest Group (NE-LRIG) had a nice meeting tonight over at Astra Zeneca's beautiful Waltham campus (woodsy borders, with a view of the nearby reservoir). The meeting was sponsored by Ion Torrent & they gave one of the three talks. All three talks were quite good, with my friend & former Millennium colleague Sunita Badola from Amgen leading off with 454 amplicon sequencing in oncology clinical trials, followed by Mark DePristo from the Broad Institute talking about the 1K genomes effort and finally Jason Affourtit from Ion Torrent (he oversees all their field applications scientists) about the Ion platform. The Ion talk gave a bit of the standard overview, followed by some slides summarizing the talks at Marco Island.

Now, when I've run previous items on Ion Torrent they have garnered a lot of comments. Some of these are positive, others (and some of my posts) not so much, with some of the commenters most charitably described as downright cynical. I won't have any data of my own for at least a month (and a machine on my own is still no more than a great desire), but I must say that if Ion Torrent is all smoke and mirrors, as some comments have insinuated, they have an awful lot of good people in on the ruse. A friend of mine who is a very experienced genomics lab head was raving about her new machine and I happened to talk to another site today and their first four runs have all come in with greater than 2X the number of reads in the specification and very good quality. About the only thing I've heard that is less than raving is that the quality drops near the end of the reads in a way that the effective read length is sometimes more like 80-90 rather than 100. Still, if you know this going in you can adapt to it.

A star before and after the talks was a PGM which was available for viewing, along with 314 and 316 chips being passed around (which I was sorely tempted to have disappear into my shirt pocket, but morals prevailed). These, for example, brought home the fact which I hadn't appreciated before that the 316 has significantly more actual surface area than the 314 chip, though it fits in the same carrier. The difference between the chips is quite visible in your hand. The 316 would appear to essentially max out the form factor, so the 318 can't keep up this trend. [corrected 3/25 per Rick's catching the typo]

Something that was emphasized tonight that I hadn't appreciated before is that there are no pumps in the PGM. Reagents are propelled by argon gas pressure, managed by valves which are themselves actuated by the argon (some electrical widgets ultimately control the valves). Also, the case apparently encompasses a lot of empty space (the Ion folks were open about this, though the machine was not open to view the innards). Presumably some of this was a conservative design leaving space for late additions (or perhaps the server), but some had to do with wanting a visually striking design.

Since the PGM instrument itself has little to do with performance of the instrument, there isn't a need to redesign it to address sequencing performance. However, there might be other reasons. While it is a relatively small benchtop instrument, space is often at a premium. A group of us from Infinity visited an academic site nearby today, and their lab made ours look spacious -- every square inch of bench was crammed with machines.

Furthermore, seeing the machine in person made it clear that in placing it, a lab must leave some space on the right side to access the wash and waste bottles along that side. Hence, if you really wanted to cram a lab the effective footprint of the instrument is a bit larger.
So, to engage in rank speculation utterly uninformed by any hard facts, I might imagine that a focus for a PGM2 would be an even smaller footprint. The four tubes in the front, which hold the nucleotides for sequencing, could be rearranged in a manner still artful (perhaps a diamond?) yet far more compact, enabling the side bottles to move to the front. Perhaps the screen could move down below the instrument -- or become an off-machine accessory capable of driving multiple instruments. Alternatively, perhaps the screen would be mounted behind the flowcell access hatch. This hatch on top (dark grey) for placing the flowcell also seems larger than necessary. So, if you really could combine all these, it could yield an instrument with an effective footprint about one half as wide or maybe better.

A question I didn't think to ask tonight is how sensitive is the instrument to the flowcell being level. I'm guessing (but certainly not a confident guess) that such devices mostly don't care; at these scales gravity isn't a dominant force. In that case, the hatch might be turned to open outwards, enabling a redesign to improve the ability to stack the instruments vertically.

Another obvious dimension would be a multi-flowcell instrument ala HiSeq 2000. Could most of the mechanical simplicity of the system be retained while enabling multiple flowcells to be run in parallel? That would be the key question.

Of course, the key driver for many of these would be if groups wanted multiple machines to drive very high throughput. I think there will be a market for this, but it is premature to think it has developed. And, it will be critical for such operations to have the promised emulsion PCR improvements (or replacement) which is promised.

What's On Your Cheat Sheet?

2011-03-22T00:24:00.000-04:00

After years of scribbling on a motley collection of pads, during my time at Infinity I've been nearly rigorous about using a single notebook for my notes -- seminar notes, phone numbers, to do reminders, random thoughts (even blog ideas). The book itself is a cheap permanently bound notebook from the local drugstore; I think they are less than $1 each.
The inside back cover of my notebook is titled "Useful Information", but I don't find very much there. Mostly it is a lot of conversion factors, but primarily for Imperial units that nobody every uses: gills, hogsheads, penny-weights, rods, scruples and such. Also such routinely accessed information such as the weight of a bushel of potatoes (60 pounds) or 1 barrel of flour (196 pounds) Other information include a 12x12 multiplication table, which was drilled into me over 30 years ago. For the metric system, about a third of the page is taken up giving the same series of prefixes with each unit. Another section has some Imperial to metric conversions.
It's interesting to think about what is curiously absent from the page. For example, the common measurements for kitchen work, tablespoons and teaspoons, are absent. Nowhere does a carat appear, nor conversion factors for the three different kinds of ounces (avoirdupois, troy and apothecary, for any European readers blissfully unaware of Imperial units). I've also missed a "stone", which is a unit of weight that shows up in historical novels -- perhaps it doesn't have precise definition. The weight of water is given in terms of a cubic ft being 2.48 gallons and weighing 62.425 pounds, rather than the usual "a pint's a pound the world around".
There are two odd values on that page given all this, but that's because I wrote them there. It is a handy place to stash info, so I have written down that 1 human genome = 6 pg of DNA (checking that in Wikipedia, apparently it is really closer to 7: 6.95 & 6.8 for female and male respectively) . The other odd value is 1 bp = 660 daltons.
Now, if I'm going to scribble in a few, why not add a bunch? Indeed, while Google will happily tell me there are 8 furlongs in a mile, it won't directly answer how much a human genome weighs. Nor will WolframAlpha -- it gave me information on human body weight in pounds. So, what else could I need there -- and if I were printing up a bunch what would I put there.
Some of the more useful molecular biology reagent catalogs have whole sections of such information. That is one challenge in designing such a information table; to be really useful it must be packed with information but at a density allowing high readability. Plus, while the catalogs use many pages, I'm trying to cram it into 1 or maybe a few (the inside cover has an equally useless class schedule grid, useless to me that is). Should I only put in what I truly can't remember, or also the things I don't have nailed so well that I can reproduce them quickly and confidently?
So, here are my current candidates, some for me and some if I were going to try to make a generally useful one. Of course, a lot of what is valuable for ready reference depends on what you are doing. At Codon I had a sheet taped by my desk with the sites for the restriction enzymes I used the most. If you have a favorite vector, the polylinker map is a useful reference. On the other hand, Planck's constant is a really important number, but one I've never needed to use in biology. So I wouldn't bother using space on it.

IUPAC ambiguity codes for nucleotides. Most I know by heart (or figure out quickly; the codes for 3 nucleotides are near the one letter they leave out), but M & K have always been a challenge. As part of cramming for this post, I now have a mneumonic that works for me: M is Methyl, for A and C, which are capable of being methylated (I think the mnemonic is supposed to be on the native structure, but I don't know that well enough). K is now the other two.
Amino acid single letter codes. I don't need this, but for a mass produced one it would make sense.
The genetic code -- without trying, I have actually memorized this, but I'm not very fast working purely from memory nor am I always confident (which is why I'm not fast)
SI prefixes in order. Again, I know most of these until you get to the two extremes, but usually have to rattle them off in order (milli=-3, micro=-6, nano=-9, pico=-12, etc).
Powers of 2. For up to 2^12, I can rattle these out. Higher sometimes comes in handy.
Tm calculation estimation using G+C and A+T counts. I don't use this often & don't really trust it, but for ballparking a Tm it might be worth having around
1 mm^3 = 1 uL and 1000 um^3=1 pL. Useful little conversions I found when I was exploring emPCR stuff (should I also put the formula for volume of a sphere in there, since I initially wrote it out incorrectly in that post?)

.That doesn't seem like nearly enough to fill up the page, but perhaps that's a good thing. I probably don't know what else it would be useful to have there, so blank space to scribble more isn't a bad idea.

My Noisy Neighbors

2011-03-18T22:56:00.000-04:00

My neighbors are up to their loud antics again; a really wild singles party. Fact of the matter is, I checked very carefully one evening before we bght the place to make sure the sound levels were what I was looking for.

Luckily, I'm not talking about a frat house or a heavy metal bar. A neighboring property has a vernal pool (a body of water which dries out in early summer, and hence cannot support fish) and the spring peepers (aka chorus frogs) tuned up for the first time of the season. Along with visiting a sugar house to see (and smell!) maple sugar being made, it's my favorite part of spring in New England.

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What will be the last Sanger genome?

2011-03-08T06:53:00.002-05:00

Even when I was finishing up as a graduate student, and only a few bacterial genomes had been published, one would periodically hear open speculation as to when the top journals would quit accepting genome sequencing papers. The thought was that once the novelty wore off, a genome would need to be increasingly difficult or have some very odd biology to keep getting in Science or Nature or such.

Happily, that still hasn't happened and genome sequencing papers still show up in the whole range of journals. I don't claim I scan every one, but I do try to poke around in a lot of the eukaryotic papers (I long since gave up on bacterial; happily they have become essentially uncountable). Two recent genomes in major journals, Daphnia (water flea) in Science and Pongo (orangutan, not dalmatian!) in Nature show that the limit has not yet been reached. These papers share another thread: both genomes were sequenced using fluorescent capillary Sanger sequencing.

Sanger, of course, was the backbone of genome projects until only very recently. Even in the last few years, only a few large genomes have been initially published using second generation technologies Read more »

Emulsion PCR: First Notes

2011-03-02T23:17:00.001-05:00

One theme in some of the comments on my Ion Torrent commentary has been around the limitations of emulsion PCR. Some have made rather bold (and negative) predictions, such as Ion Torrent dooming themselves to short read lengths or users being unable to process many samples in parallel without cross-contamination.

Reading these really drove home to me that I didn't understand emulsion PCR. I've done regular PCR (once in a hotel ballroom, of all places!) but not emulsions. It seems simple in theory, but what goes on in practice? My main reasoning was based on the fact that emPCR is the prep method for both 454 and SOLiD; 454 clearly demonstrates the ability to execute long reads (occasionally hitting nearly a kilobase) and SOLiD the ability to generate enormous numbers of beads through emPCR. I also have a passing familiarity with RainDance's technology (we participated in the RainDance & Expression Analysis Cancer Grant program). But, I've also seen a 454 experiment go awry in a manner which was blamed on emPCR -- a small fraction of primer dimers in our input material became a painful fraction of the sequencing reads. Plus, there is that temptation to enter the Life Tech grand challenge on sample prep, or attempt to goad some friends into entering. So it is really past time to get more serious about understanding the technology.

So, off to the electronic library. Maddeningly, many of the authors in the field tend to publish in journals that I have less than facile access to, but between library visits, PubMed Central and those that are in more accessible journals, I've found a decent start.
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When Will Life Technologies Get Serious About Their Grand Challenges?

2011-03-01T23:24:00.000-05:00

My recent run of posts on Ion Torrent certainly garnered a lot of comments, and it would be much less than honest to say that many of the comments were far less favorable to Ion Torrent than what I have written. Indeed, many were not terribly favorable on me given what I had written about Ion Torrent -- one even asked if I "felt used" as part of a publicity stunt. (BTW, I don't -- if I can't ask the hard questions I have nobody to blame but myself).

One Ion's other very public events around the Ion Torrent has been to announce a series of three challenges to improve the performance of the instrument system (a fourth has been announced centered around SOLiD and three others have yet to be unveiled). The winner of a challenge can get $1M in prize money.

Now, contests along these lines have been successfully used by companies and organizations to drive technologies forwards. Netflix successfully crowdsourced better prediction of a user's movie tastes. The most spectacular success for such a contest was the winning entry for the Ansari X-Prize, SpaceShip One. Google is currently sponsoring a contest to land a rover on the moon and transmit HDTV images, which I look forward to eagerly.

Unfortunately, so far Life Technologies & Ion Torrent's contest seems to be all hat and no cattle. While the three goals have been announced (double the output per run, halve the sample prep time and double the accuracy), nothing else is in place. Each competition is apparently separate; there's no prize for halfway success on two of the axes. If they are serious about attracting competitors, they need to get down to brass tacks.

Now, I can't say I'm surprised. Not only has Ion shown a penchant for loudly trumpeting their progress prior to demonstrating it, but in their previous contest showed a certain degree of haste and a few punchlist items. In the first contest, submissions for how to use the instrument were judged to yield two U.S. winners (followed recently by two European winners). Each submission consisted of two parts; in the original rules it wasn't clearly stated what the distinction was between the two parts (perhaps it should have been obvious, but I don't routinely write grants) other than the rules stated a word limit for one of them. Once you tried to submit, however, then the word limit on the second section became apparent. Ion also ended up extending the deadline for submissions, which can either be seen as generous or irritating -- in the latter case, if you've burned midnight oil & spent part of a vacation chopping down an overlong second section to get your entry in on time. Importantly, that contest has a tiny fraction of complexity of any one of these contests.

Starting with, what are the rules? One key question will be around cost. For example, can a winning entry for sample prep use an instrument that costs much more than the PGM? That's not an absurd concept. Can the double the output prize be won by a sample prep process that takes a long time? For example, can I assay to find only DNA-bearing beads & then use a micromanipulator to position them? That is obviously a deliberately absurd proposal. But, unless the rules are carefully crafted someone will attempt a silly entry, and Ion will have a real mess if they are forced to put the laurels on silly.

A key & challenging area is around intellectual property (IP). The first obvious issue in this department is how much IP can you retain when submitting an entry? Obviously Ion isn't interested in paying out $1M to something they can't use -- so is the $1M in effect a license fee (with no royalties?)? On the other side of the IP coin, how much IP can a winning submission use which the submitter does not have rights to? For example, some wag might submit a sample prep protocol that is bridge PCR using in part Illumina reagents. But more complicated would be methods that only an IP lawyer can decide either infringe or build on some prior patent. If it's Life's patent, presumably they wouldn't care -- but an Illumina or Affy patent would be an entirely different fish.

Materials are going to be another critical issue issue for the yield and sample prep challenges. Any reasonable scheme for attacking these is going to get very expensive if complete kits must be purchased each time. For example, you may want to hammer on the beads without ever actually putting them on a chip. Will Ion give at-cost access to the specialized reagents (such as beads)? Furthermore, how much information are they willing to give out on the precise specs. For example, suppose a concept requires attaching something different to the beads than standard -- will specifications be provided to create appropriate beads?

Another key question is what samples? Will Life Tech supply the samples to use for improving yield or does a group get to define them? A devious approach to winning the prize would be to develop a sample which preps very badly with the standard prep. An attempt could be made to legislate this possibility away, but there would be significant advantage to standardized samples. But should these be real world samples, idealized samples (such as a pure population of a single fragment) or deliberately hard real world situations (e.g. an amplicon sample with a high fraction of primer dimers)? In a similar vein, what dataset will be made available for the accuracy challenge?

Now, Life is promising more information this Spring, and since that is still a few weeks away (or do they go by Punxsutawney Phil?). I really hope in the future they try to hold back their announcements until they're really ready to go. It doesn't help that the Twitter rehash scrolling on their screen is full of links that might provide more information, but none of them work. They really need to rethink their strategy of piecemeal delivery that can do nothing but frustrate the possible entrants in the contest.

Part of my frustration is I can't help but ponder throwing my hat in the ring. It's not hard to think of ideas for the sample prep problem and while I couldn't do the experiments I do have friends who could (time to get the core Codon team back in action!). Of coures, working out the IP headache would be an issue (unless the work was done at work, which is sadly too far afield of what we do to be a responsible course). I can also imagine a number of academic groups and even a few companies which might seriously consider entering some of these challenges. I'd love to talk up the accuracy challenge with computer geeks I know. The problems are of a very attractive sort for me -- you can very quickly generate very large and rich datasets, enabling quantitative approaches (such as Design of Experiments) to optimization. A lot of data can be generated without actually running chips but using even lower cost methods (such as microscopy or flow cytometry) to measure key aspects. But with nothing concrete to point at, it seems rather pointless to start scheming.

But, while I can't actually move forward on any of these, I can do a bit of homework on emulsion PCR. I'll try to write up that homework later this week, as it's been informative to me and I believe puts me in a better position to handicap Ion Torrent's claims on sample prep -- and various comments on emPCR from the previous posts.

Reflecting & Responding on Today's Comments

2011-02-24T23:40:00.000-05:00

It's nice to see my piece on the Ion Torrent 318 chip get a lot of comments, as well as being tweeted and some additional comments at SEQAnswers. I love getting comments, especially those that constructively challenge my analysis.

Two commenters threw some cold water on my enthusiasm, pointing out that the 318 is not scheduled to be released until September, and then only to early access customers. I'll confess to not asking careful enough questions on the timing & letting my desires get ahead of me. Ion Torrent will need to deliver these chips on time and working at specifications in order for scientists to believe that there really will be the promised regular upgrades. MiSeq is vaporware at the moment also, but Illumina has a strong track record for rolling out instruments.

One commenter took me to task for suggesting that Ion Torrent will simply roll right over Illumina. I don't mean to suggest this will happen, only that Ion Torrent may offer a very stiff challenge. If the PGM chip upgrades, sample prep improvements and read length improvements come as promised, then the PGM may well march up through Illumina's product line. That's not to say Illumina will remain static; they have clearly demonstrated great skill at pushing their platform to amazing abilities. The MiSeq may offer a new path to greater performance; there has already been speculation over at SEQAnswers on possible paths forward. For example, will the faster cycle times mean an opportunity for longer read lengths (if the reagents are not stable over very long runs)? Will Illumina roll out a multi-flowcell version of the MiSeq, enabling very high-throughput rapid-turnaround sequencing? I'd put more money on the latter than the former. Another thread has surfaced rumors that a HiScan upgrade is imminent which will give it HiSeq-like capabilities; for groups running both arrays and sequencing this could be useful. How much denser can Illumina pack the flowcells? Only time will tell. But, as I pointed out before, longer reads will have limited impact on overall throughput if the two reads are overlapping (though accuracy may be improved).

On the other hand, I do see as excessive cynicism (bordering on editing the truth) that AGBT had only Ion Torrent talking about Ion Torrent; given that two talks (Bolen from NCI and Nusbaum from the Broad; the Bolen talk will apparently be at ABRF as well) described actual experiences. Granted, neither was really independent of the company. It is critical for Ion Torrent's reputation that data start appearing in public spaces, especially data from regular customers.

Ditto for the experimental protocols; one commenter had a question about amplicon sequencing which will be important for Ion Torrent's success. The sooner applications are adapted to the system, the sooner customers will be buying machines and chips. Of course, a great strategy IMHO would be to put some sequencers in the hands of genomics bloggers.

As far as the question of reads per chip increasing faster than the number of sensors per chip, my understanding is that Ion Torrent expects that better sensor loading and utilization. Again, how vaporous is this? That's hard to tell -- and why feedback from actual users is desperately needed.

On the question of amplicon and read length for Ion Torrent, those would seem to be different fish (though I don't have direct experience). Amplicon length will be a matter of the emPCR conditions. I'm unaware of any signal amplification in the 454 chemistry -- I thought the signal cascade was strictly linear. But, again that is outside my expertise and I would enjoy being educated on the subject.

With regard to whether emPCR can be converted from a liability to a non-issue, it's reasonable to be skeptical. But, it isn't obvious why emPCR can't be compressed in time (though I haven't done emPCR, so that's a stretch for me to comment on). In my discussion with them, they pointed to cutting the number of cycles as one time savings. In any case, how hard as anyone tried to really optimize emPCR for speed? Only on the 454 might it have been a serious concern, and the cost of a 454 run is not going to encourage many folks to push for many runs a day. On the other hand, I don't see it as "disingenuous" the comparison I made; I've been at a company that had techs running PCR (again, conventional not emPCR) day in and day out, and any company that runs production Sanger is in a similar boat. Indeed, any big SOLiD or 454 shop must do this as well. What will the nature of the new emPCR kit be? I have some speculations, but that's a whole 'nother post.

One other surprise today from Ion Torrent is the announcement that they have sequenced the genome of Intel co-founder Gordon Moore. One claim from this is that the coverage was more uniform than with other platforms. Obviously, either publication or release of the data is needed to vet this claim. Even if they used 318 chips, this was an expensive demonstration project (I don't know the actual coverage; I don't have a subscription to In Sequence) -- even at 10X coverage it would be $20K (retail) in chips. Apparently this was an exclusive for GenomeWeb's In Sequence, as Ion doesn't even seem to have a press release out. I hope they plan to publish this in a journal soon, or if not make the primary data available for further study.

Has Ion Torrent Taken A 318-Sized Lead over MiSeq?

2011-02-23T10:02:00.003-05:00

About a week ago, Ion Torrent's President Greg Fergus and Head of Marketing Manesh Jain were kind enough to engage me in a nearly an hour of discussion about the Ion Torrent platform. One agreement prior to our discussion is that I would withhold this piece until their announcement today of the 318 chip for the system (I also volunteered to let them see a draft of this in advance to ensure I had not misrepresented anything).

A key theme on their side is a certain degree of feeling that the wrong questions are being asked in the analysis of PGM versus MiSeq -- and an eagerness to shift the discussion. They wished to emphasize a number of points, and after the discussion I can see the validity of many of these.
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Is Being Lucky the Same as Being Smart?

2011-02-23T00:19:00.000-05:00

Forbes has an article co-written by Matthew Herper and Robert Langreth titled "The Next Big Move For The Smartest Biotech Investor", profiling Randal Kirk. Kirk is described as one of the few billionaires who can ascribe that status to biotechnology. Kirk made his money through two companies in the psychiatric drug space: New River Pharmaceuticals developed an ADHD drug (lisdexamfetamine) and then was acquired by Shire whereas Clinical Data developed an antidepressant (vilazodone) and was then purchased by Forest. A key part of the article profiles Kirk's investment in a little-known and secretive synthetic biology company called "Intrexon".

The title is probably meant to gall; it certainly raises my hackles. The most obvious quibble is that it isn't clear either of the drug development companies were really biotech. Of course, that would require defining biotech, but it ideally it would refer to companies which highly depend on recombinant DNA and related technologies. Now, such technologies are embedded in virtually all drug development today, but neither of these drugs sounds like they used much. Both drugs are interesting twists on prior approaches (though I'm not enough of a chemist to judge the novelty of vilazodone).
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Will Cheap Gene Synthesis Squelch Cheaper Gene Synthesis?

2011-02-20T13:07:00.002-05:00

Among the vast piles of items which I've meant to write about but have slipped are a paper last year on gene synthesis and some subsequent announcements about trying to commercialize the method described in that paper. This is an area in which I have past experience, though I would never claim this gives me indisputable authority or omniscience in the matter.

The paper, primarily by scientists from Febit but also with two scientists from Stanford and George Church in the author list, finally describes an interesting approach to dealing with some serious challenges in gene synthesis which substantially increase the costs. By finally, I mean that the idea has certainly been kicking around for a while and was mentioned when I visited Codon Devices in the fall of 2006 looking for employment.

To fill in some background first, gene synthesis is a powerful way to generate DNA constructs which can enable all sorts of experiments. The challenge is that the cost of gene synthesis, currently starting at around $0.40 per base pair for very easy and short stuff (say, less than 2Kb), tends to restrict what you can use it for. I have a project concept right now that would be a slam dunk for gene synthesis -- but not at $0.40/bp (which I think I couldn't even get for the project). Whack that price by a few factors of two and the project becomes reasonable.

There are many cost components to commercial gene synthesis, and only someone who has carefully looked over the books while wearing a green eyeshade is going to have a proper handle on them. But three of the big expenses are the oligos themselves, the sequencing of constructs to find the correct ones and labor. What the Febit paper does is illustrate a nice way to tackle the first two in a manner that shouldn't require a lot of labor.

The oligo cost is a serious issue. Conventional oligos can be had for around $0.08 or maybe a bit less a base. However, each base in the final construct requires close to 2 bases in the oligo set. Some design strategies might get this down a bit. However, conventional columns generate far more oligo than you actually need. An approach which has been published (but not commercialized as far as I know), is to scale down the synthesis using microfluidics. This method matches better the amount synthesized and the amount you need, though the length and quality of the oligos needs refinements from what was reported in order to be truly useful. Microarrays are a means to synthesize huge numbers of oligos, but their quality also tends to be low and the quantity of each oligo species is much too small without further amplification. Amplification schemes have been worked out, but add to the processing costs of the oligos.

What Febit and company have done is take those microarray-build oligos and screen them using 454 sequencing. The beads containing the amplicons with correct oligos are then plucked out of the 454 flowcell (with 90% success of getting the right bead) and used as starting points.

Now, this has several interesting angles. First, it has been challenging to marry the non-Sanger new sequencing technologies to gene synthesis. The new technologies tend to have short reads, too short to read even a short construct. The new technologies also require library construction and it is difficult to trace a given sequence back to a specific input DNA. In other words, short read technologies are great at reading populations, but not individual wells in a gene synthesis output. Sanger on the other hand, is ill-suited for populations but great for individual clones. One solution to this problem is clever pooling and barcoding strategies, but these necessitate having enough different clones to be worth pooling and barcoding. In other words, second generation sequencing is difficult to adapt to retail gene synthesis, but looks practical for wholesale gene synthesis.

Getting the oligos right has important positive side-effects. While the stitching together of oligos into larger fragments (and larger fragments into still larger ones) can generate errors, and awful lot of the problems stem from bad input oligos. Not only can error rates be troublesome, but some of the erroneous sequences may have advantages over the correct ones in later steps. For example, a deleted fragment may PCR more efficiently than the full length, and slightly toxic gene products may be disfavored in cloning steps over frameshifted versions of the same reading frame. So, by putting the sequencing up front it should be possible to reduce the later sequencing downstream. So even if that sequencing remains Sanger, it should be possible to do a lot less.

Okay, that's the science. Now some worries about the business. Febit announced in January they are looking for investors to fire off a new company to commercialize this approach. This makes good business sense, since Febit itself must be encrusted with all sorts of business barnacles, having lurched from one business to another in trying to commercialize their microfluidic microarray system. Previously failed attempts include gene synthesis as well as microarray expression analysis and hybridization capture (I even ran one experiment with their system, whose results certainly didn't argue for them staying in that business!). The press release stated they were hoping to attain pricing in the 0.08$ per base range, which would make my current experiment concept feasible. That would be great.

Now, they will need to refine their system and perhaps adapt other sequencers. A 454 Jr would probably not be a difficult adaptation, but moving on to Ion Torrent must be tempting. Getting things to work for one paper and one set of genes is unfortunately different than being able to keep things working over an entire spectrum of customer designs.

Which leads me to where I think they will have a great challenge, though one which I think can be finessed with the proper business approach. They will be brining to market a methodology whose benefit is cost at the expense but with the caveat of attaining that cost advantage only with sufficient volume. Initially, they will be unable to reliably predict delivery times (due to kinks showing up). Finally, they are adding some additional processing steps (454 sequencing, bead recovery & oligo recovery from the bead) which may add to the time.

The abyss into which this new company must plunge is a world in which very fast gene synthesis is available from a large number of vendors in the $0.40 price range. So, they must find very large customers who are willing to be a bit patient and keep their pipeline filled. Such customers do exist, but they aren't always easy to find and pry away from their existing suppliers. In theory much cheaper synthesis would unleash new orders for projects (such as mine) which are too costly at current prices, but that is always a risky assumption to bank a company on (c.f. Codon Devices' gene synthesis business).

It's the alternative route that I predict this NewCo is likely to go down. That would be to link up with an established provider in the field. Said provider, through their salespersons and sales software, could offer each customer an option -- I can build your genes for $0.40 if you want them fast or hack that down to $0.10 a base if you can wait. In order to preserve customer satisfaction, that long time would need to include an insurance period to build the genes by the conventional route if the new route fails -- but of course if you are frequently forced to build $0.40/bp genes for which you charged $0.10/bp, that would be financial suicide.

So, in summary, I think this is a clever idea which needs to be pushed forward. But, after a long gestation in the lab, it faces a very rocky future in the production world. I hope they succeed, because it is not hard to imagine projects I would like to do which would be enabled by such a capability.

Roads That Really Need Taking

2011-02-17T23:26:00.003-05:00

There is a provocative Perspective piece in a recent Nature titled "Too many roads not taken". In it, the authors summarize some more extensive work (posted on Arxiv) and others showing that in several major fields of protein research, the biological community seems to be stuck on a few old favorites, with insufficient attention to other key players.

In the Nature piece, the focus is on protein kinases and nuclear hormone receptors, both key druggable classes. In the kinase area, they have a graph (the Arxiv site leads to the actual data) showing that the graph of attention (measured in bibliographic citations) given to kinases prior to 2003 is generally predictive of the attention given them in 2009; only a handful of kinases with little attention in the first time period are hot areas now. Of course, if the rest of the kinome were boring this would be justifiable, but they point to several papers which indicate that many of the unstudied kinases (for example, this series of shRNA experiments)

A very striking conclusion is underlined by their plot of NHR citations by gene, which is expanded upon significantly in the Arxiv preprint. Their contention is that the availability of chemical probes drives analysis in this family, and in their rank-ordered histogram of activity the proteins with chemical ligands are all at the top of the list. Granted, at the breakpoint the bars are nearly equally tiny on either side of the divide, but it does suggest a strong correlation. Easy to build universal approaches such as RNAi are very powerful, but especially for any kind of in vivo study are suboptimal -- not only are they hard to deliver, but utterly wiping out a protein is not a subtle way to probe a system. The Arxiv piece goes into more detail and posits that you really need good compounds, ones with good bioavailability and such. In other words, drug-like compounds.

So, they plead that we need to invest more in getting such compounds into the research community. Some of this could take the form of funding screening efforts in the public sphere, which like sequencing the human genome isn't the sexiest project from a scientific perspective -- just like the foundation is never the sexiest part of a great building. But foundations matter!

An alternative, suggested by the Arxiv paper, is that compounds may exist in the patent literature but are not widely commercially available. So, a public effort might focus on mining the patent literature, synthesizing various claimed compounds and verifying their activity. Of course, some (many?) compounds in the patents might not stand up to scrutiny. Another alternative would be to attempt to devise incentives for industry to release some of the inhibitors they have in house; obviously this would be a challenge as such compounds might prove to be the starting points for valuable drugs -- but of course they never will until someone proves the underlying kinases valueable. Some companies have generated unbelievable databases on kinase inhibitors (for example, this recent study) which have the potential to fill in the missing spots on the kinase tool compound grid.

I can't help thinking of the new translational medicine institute going in at the NIH. Trying to move research findings into the clinic to benefit patients is hard to argue against, but perhaps the NIH needs to also pour some resources into the humble task of finding basic tools.

Wrapping up AGBT

2011-02-08T23:26:00.002-05:00

AGBT ended on Saturday, but both Saturday & Sunday were crammed with personal stuff. Monday night was some down time, so now tidying up.

Anthony Fejes has a nice final post on the meeting (along with all his useful notes); it's also interesting to see what a boost in traffic he gets for AGBT (and well deserved!). One very positive Twitter note: one presenter personally thanked him for blogging his talk; I know I get a thrill whenever authors visit this space with comments & it's good to see that sometimes blogging really is opening up a two-way conversation. A more constrained view on the proceedings can be seen in an investment house's research note.

A few last impressions of my own based on Anthony's notes and Twitter. Nanopores seem to be making progress -- but still quite distant from being able to generate any data. PacBio appears to have faced reality and is positioning themselves in areas where their strengths (read length, speed) are strengths and accepting that their per-base quality isn't going to be adequate for many applications. Metagenomics is something I really need to do a personal deep dive into -- it's often sounded cool & I'd love to know more. The whole idea that each region of my skin has a different flora is pretty amazing -- and one wonders what the scale of changes are (indeed, I wonder about how different my scales are than adjacent area -- it's winter and I have several serious dry patches). The Broad is using 500Gb of RAM on their machines doing de novo vertebrate assembly; given that my first and second computers came with 1Kb of RAM, that's pretty mind blowing.

Someone asked for my Twitter table. Alas, I didn't save some intermediate results & Twitter's search mechanism apparently now won't go back beyond Thursday night, but below is what I can spit out -- with the URLs turned into hyperlinks. Maybe before the next meeting I'll engage in some lily-gilding and have this thing build a SQLite database of results. Twitterstatistical summaries for the whole meeting are a bit interesting.

Tue, 08 Feb 2011 21:30:16 +0000 KamounLab http://bit.ly/dXiQXS￢ﾀﾝ AGBT Excellent summary: ￢ﾀﾜ@genomeresearch: What investors heard at - summary & analysis from A Murphy of William Blair
Tue, 08 Feb 2011 18:57:00 +0000 genomicslawyer http://bit.ly/dXiQXS AGBT What investors heard at - summary & analysis from Amanda Murphy of William Blair:
Tue, 08 Feb 2011 12:07:24 +0000 samuellampa HTSeq,NextGenSeq,AGBT Or maybe is more time-proof (and shorter) than ...
Tue, 08 Feb 2011 12:05:30 +0000 samuellampa sequencing,NextGenSeq,AGBT (Seems clashes with some music production stuff etc ... )
Tue, 08 Feb 2011 12:04:43 +0000 samuellampa NextGenSeq #AGBT Folks, doesn't need a better hash tag? Some brainstorming? ...or what about the one above?
Mon, 07 Feb 2011 23:03:32 +0000 apfejes AGBT @PeroMHC is always at the same place on Marco Island.
Mon, 07 Feb 2011 22:47:05 +0000 PeroMHC AGBT Where is 2012??
Mon, 07 Feb 2011 22:16:49 +0000 finchtalk AGBT #1kgenome and now a chat Cliff Reed and Eric Schadt. From to Seattle. Luke Timmerman will moderate.
Mon, 07 Feb 2011 20:10:54 +0000 apfejes AGBT @drio Was it George Vacek, poster 214 in the book?
Mon, 07 Feb 2011 19:52:04 +0000 drio #AGBT does anyone remember the name of the company that had the poster about using FPGAs with velvet? or the person that presented.
Mon, 07 Feb 2011 18:23:55 +0000 bachinsky agbt @apfejes Thanks you for tweets
Mon, 07 Feb 2011 17:44:09 +0000 drjonboyg AGBT @apfejes thanks for writing up the sessions, your blog is a great resource.
Mon, 07 Feb 2011 17:39:10 +0000 apfejes http://blog.fejes.ca/?p=363 AGBT Final thoughts and summary of this year's conference. http://blog.fejes.ca/?p=363
Mon, 07 Feb 2011 16:53:24 +0000 JVJAI http://blog.fejes.ca AGBT @apfejes Thank you so much for your tweets & great meeting notes at
Mon, 07 Feb 2011 16:44:34 +0000 apfejes AGBT You're all welcome for the conf notes, @ntuseem, @ChaunceyGrattan, @genomeresearch, etc! I'm glad you're finding them to be useful!
Mon, 07 Feb 2011 16:31:58 +0000 PMedPartners http://summarizr.labs.eduserv.org.uk/?hashtag=agbt agbt Check out the @twapperkeeper summary: 272 tweeters! http://summarizr.labs.eduserv.org.uk/?hashtag=agbt
Mon, 07 Feb 2011 16:05:26 +0000 j3moore AGBT? @drjonboyg @djschlesinger: would you mind sharing what you found so impressive about the BioNanomatrix talk at
Mon, 07 Feb 2011 15:46:44 +0000 genomeresearch AGBT Genome Research back in Cold Spring Harbor. Thanks to all tweeps esp @apfejes @deannachurch @djschlesinger @drjonboyg @girlscientist
Mon, 07 Feb 2011 15:44:04 +0000 finchtalk http://yfrog.com/h7baknp #AGBT - once again, great conf. See photo for my summary.
Mon, 07 Feb 2011 13:02:10 +0000 deannachurch AGBT @drjonboyg thanks! And thanks to all of the tweeps for great feedback.
Mon, 07 Feb 2011 11:13:16 +0000 drjonboyg AGBT On reflection, a big thanks to @deannachurch and the other organizers, it was an excellent and informative meeting.
Mon, 07 Feb 2011 01:25:21 +0000 girlscientist AGBT Have to give recap of for lab mtg tomorrow. Now going to do it with lighted box on head.
Mon, 07 Feb 2011 00:47:11 +0000 girlscientist AGBT Back home -- thanks tweeps, it was a lot of fun. But snow forecast here tomorrow?!? Take me back to the beach....
Sun, 06 Feb 2011 21:21:38 +0000 deannachurch http://yfrog.com/gyhq5cj agbt,hoedown new in Roche sales.
Sun, 06 Feb 2011 19:25:09 +0000 ChaunceyGrattan AGBT @apfejes many thanks for the constant information stream. Future conf attendees should take note of how its done.
Sun, 06 Feb 2011 14:19:26 +0000 deannachurch AGBT,detox,sleep Bye-bye see you next year.
Sun, 06 Feb 2011 13:50:12 +0000 apfejes AGBT Apologies to Dr. Mungall, who's name has now been mispelled with one l all over the internet because I can't spell!
Sun, 06 Feb 2011 01:36:25 +0000 djschlesinger AGBT Are fish tacos traditional cowboy food?
Sun, 06 Feb 2011 01:28:56 +0000 omespeak http://yfrog.com/h46w8cyj AGBT The final swag: cowboy hats and bandanas at the dinner.
Sun, 06 Feb 2011 00:46:34 +0000 rforsberg http://cphx.org/ AGBT Thanks for a great conference, see you next year at or see you at Copenhagenomics in June !! -
Sat, 05 Feb 2011 23:00:16 +0000 SOLiDSequencing http://bit.ly/solid_community sequencing #AGBT folks: want to stay up-to-date on @SOLiDSequencing news? Join the SOLiD Community:
Sat, 05 Feb 2011 22:54:34 +0000 SOLiDSequencing http://bit.ly/pgm_winner AGBT?,PGM Missed @LifeCorporation Triathlon Get the recap: Congrats to @iontorrent winner!
Sat, 05 Feb 2011 21:41:54 +0000 phylogenomics AGBT,sarcasm Basking in my brilliance for starting to track meeting during last talk
Sat, 05 Feb 2011 21:38:11 +0000 deannachurch AGBT Discussion on how people want to see improve. Suggestions?
Sat, 05 Feb 2011 21:37:28 +0000 rdeborja #AGBT last talk is now over. Met lots of people in the cancer genomics field.
Sat, 05 Feb 2011 21:37:15 +0000 phylogenomics http://ff.im/-xpHIb AGBT,Experiment Alternative Real Time Twitter Feed for
Sat, 05 Feb 2011 21:37:01 +0000 djschlesinger AGBT Amazing meeting. Many thanks to the organizers. Only suggestion: would like to have had more time for posters.
Sat, 05 Feb 2011 21:35:01 +0000 apfejes AGBT After this talk, maybe Ellen Wright Clayton was right - you can go too far with sequencing!
Sat, 05 Feb 2011 21:32:17 +0000 girlscientist AGBT Schadt suggests we should all hook up PacBio machine to efflux from toilets so we can have health report on house occupants.
Sat, 05 Feb 2011 21:31:59 +0000 dsexton_2 agbt that may have been the scariest scenario ever presented
Sat, 05 Feb 2011 21:30:44 +0000 apfejes AGBT Waiting for announcement that Pac bio has collaborated with google to sequence your DNA as google maps & photographs you...
Sat, 05 Feb 2011 21:28:36 +0000 djschlesinger AGBT Pacific Biosciences SMRT + Illumina SBS = some damn cool data
Sat, 05 Feb 2011 21:27:41 +0000 deannachurch AGBT ES: having success combining Illumina and PacBio data.
Sat, 05 Feb 2011 21:26:32 +0000 phylogenomics http://ff.im/-xpGpv AGBT Tracking Advances in Genome Biology & Technology meeting w/ Twitter Widget:
Sat, 05 Feb 2011 21:25:14 +0000 deannachurch AGBT ES: Profiling sewage as a proof of principle, because "Everybody poops".
Sat, 05 Feb 2011 21:22:30 +0000 girlscientist AGBT If you see someone in the airport bathroom looking crazy, it's just Eric Schadt sampling toilet flush handles for sequencing.
Sat, 05 Feb 2011 21:21:53 +0000 girlscientist AGBT Schadt: want to develop real-time disease weather map by viral metagenome sequencing of travel centers etc.
Sat, 05 Feb 2011 21:21:21 +0000 deannachurch AGBT ES: "Disease weather map" surveying viral metagenomes to try to predict outbreaks.
Sat, 05 Feb 2011 21:19:43 +0000 djschlesinger AGBT￢ﾀﾝ Has he taken a breath yet? ￢ﾀﾜ@girlscientist: @apfejes no one, no one can keep up with Eric Schadt. You have done a great job.
Sat, 05 Feb 2011 21:19:16 +0000 deannachurch rapidfire,sorry,AGBT ES: Maybe the 93 contigs was just from Illumina/454- not sure-
Sat, 05 Feb 2011 21:18:02 +0000 deannachurch AGBT ES: Finishing and layering in Pac Bio- complete genome. 8 contigs >1Kb, 6 contigs covering 99% of genome.
Sat, 05 Feb 2011 21:17:40 +0000 girlscientist AGBT @apfejes no one, no one can keep up with Eric Schadt. You have done a great job.
Sat, 05 Feb 2011 21:17:06 +0000 apfejes finallydefeated,AGBT Cool talk from Pac Bio, but I just can't blog fast enough to keep up with the firehose of data presented.
Sat, 05 Feb 2011 21:16:47 +0000 deannachurch AGBT ES: 244X coverage in 454, Illumina and PacBio to do an assembly: still had 93 contigs after hybrid assembly.
Sat, 05 Feb 2011 21:16:26 +0000 girlscientist AGBT Schadt et al. have gone on from NEJM paper to combine Illumina, PacBio regular, and PacBio strobed sequence reads to complete genomes.
Sat, 05 Feb 2011 21:13:55 +0000 deannachurch AGBT ES: sequence in 90 minutes, then on to the comparative genomes; "quick as serotyping or fingerprinting".
Sat, 05 Feb 2011 21:11:36 +0000 djschlesinger AGBT Eric: Cholera strain to sequence data (on PacBio) to published paper in 4 weeks!!
Sat, 05 Feb 2011 21:07:22 +0000 djschlesinger AGBT @apfejes for long term sustainability, I agree. However I think sugar cane would be ideal to fill an immediate need.
Sat, 05 Feb 2011 21:07:00 +0000 djschlesinger AGBT Eric Schadt from PacBio on rapid pathogen ID.
Sat, 05 Feb 2011 21:06:54 +0000 deannachurch AGBT ES: OK- two minutes in and very cool visualization...
Sat, 05 Feb 2011 21:05:14 +0000 deannachurch AGBT Eric Schadt to end the day: sequencing for impacting health.
Sat, 05 Feb 2011 21:05:10 +0000 girlscientist AGBT Last talk of = Eric Schadt and PacBio's cholera work.
Sat, 05 Feb 2011 21:04:29 +0000 girlscientist AGBT Zhong Wang offers ftp address for data, "if you want to stick your hand in cow rumen and see what you get out." Check JGI site.
Sat, 05 Feb 2011 21:03:36 +0000 SEQanswers agbt Ah the fistulated cow, brings back memories of UC Davis!
Sat, 05 Feb 2011 21:03:18 +0000 apfejes http://blog.fejes.ca/?p=357 AGBT Notes and thoughts on Zhong Wang's talk on cow gut microbiome + cellulases: http://blog.fejes.ca/?p=357
Sat, 05 Feb 2011 21:02:21 +0000 djschlesinger AGBT good point, was thinking biological: @mlb_house: @djschlesinginger or, how about improving solar panels and directly harvesting solar?
Sat, 05 Feb 2011 21:01:39 +0000 apfejes AGBT @djschlesinger Sugar cane still produces a lot of cellulosic waste - it's better to use the cellulose than compete for the glucose.
Sat, 05 Feb 2011 21:00:44 +0000 mlb_house AGBT @djschlesinginger or, how about improving solar panels and directly harvesting solar?
Sat, 05 Feb 2011 20:59:52 +0000 MaliciaRogue http://bit.ly/eMtbWd AGBT . ￢ﾙﾻ @genomeresearch: JGI cow rumen metagenome assembly by Velvet
Sat, 05 Feb 2011 20:59:21 +0000 deannachurch AGBT Wang: Doing single cell sequencing to try to validate assemblies from metagenomes.
Sat, 05 Feb 2011 20:57:39 +0000 genomeresearch http://bit.ly/eMtbWd AGBT JGI cow rumen metagenome assembly by Velvet
Sat, 05 Feb 2011 20:52:51 +0000 djschlesinger AGBT Instead of finding or engineering enzymes to turn cellulose to sugar, why not engineer sugar cane to grow in more temperate climates?
Sat, 05 Feb 2011 20:48:46 +0000 djschlesinger AGBT @apfejes I believe switchgrass yields more energy than corn per equal mass.
Sat, 05 Feb 2011 20:46:11 +0000 deannachurch AGBT @apfejes switchgrass is cheaper to grow- and it is not a human food source so you don't create price competition.
Sat, 05 Feb 2011 20:45:20 +0000 SOLiDSequencing PGM,AGBT Congrats to Adam English, Baylor, winner of @iontorrent sequencer. $10K ($1/total points) goes to Komen for the Cure!
Sat, 05 Feb 2011 20:45:09 +0000 apfejes AGBT Odd that switchgrass was used - it's grows fast, but corn cellulose or other grass stalks are plentiful as a wasteproduct...
Sat, 05 Feb 2011 20:41:17 +0000 deannachurch AGBT Wang: JGI has a cow with a "door" so they can efficiently sample rumen.
Sat, 05 Feb 2011 20:41:10 +0000 SOLiDSequencing AGBT,sequencing @apfejes really enjoying your live blogging (where allowed). Thanks for the great conference notes.
Sat, 05 Feb 2011 20:39:46 +0000 cslamo #AGBT folks stop RTing BS or I'll unfollow everyone!
Sat, 05 Feb 2011 20:36:22 +0000 deannachurch AGBT Zhong Wang: using genomics to address our fuel needs.
Sat, 05 Feb 2011 20:34:18 +0000 apfejes http://blog.fejes.ca/?p=353 AGBT Rather incomplete notes on Mark Akeson's (movie filled) talk on nanopores: http://blog.fejes.ca/?p=353
Sat, 05 Feb 2011 20:33:01 +0000 girlscientist AGBT Next talk: Zhong Wang, JGI, metagenomics of cow rumen. Just published in Science.
Sat, 05 Feb 2011 20:32:24 +0000 girlscientist AGBT Akeson: next steps include seq and re-seq indiv DNA templates, and generating read-lengths signif. longer than industry standards.
Sat, 05 Feb 2011 20:29:16 +0000 nanopore http://bit.ly/fg5iB1 AGBT MA: phi29 high processivity and catalysis rate, animation representing data from JACS paper
Sat, 05 Feb 2011 20:24:40 +0000 nanopore AGBT MA: movie - watching Klenow fragment add bases. Trace showing measurement of ionic current
Sat, 05 Feb 2011 20:23:52 +0000 KamounLab http://tinyurl.com/4mr8gk7 AGBT Link to Lorena Beese's movie "processive DNA synthesis in a polymerase crystal" mentioned by Mark Akeson
Sat, 05 Feb 2011 20:18:10 +0000 nanopore AGBT MA: how to control translocation of ssDNA through a nanopore..add a processive enzyme. up first Klenow fragment
Sat, 05 Feb 2011 20:16:09 +0000 girlscientist AGBT Akeson: group of Jens Gundlach doing promising work on changing over from alpha-hemolysin nanopore to MspA.
Sat, 05 Feb 2011 20:13:45 +0000 nanopore AGBT MA: Bayley group's recognition of bases inc modified bases on a strand
Sat, 05 Feb 2011 20:13:04 +0000 nanopore AGBT MA: in wild type hemolysin, many bases contribute to signal - so engineering of reader heads needed for resolution
Sat, 05 Feb 2011 20:11:42 +0000 djschlesinger AGBT Protein nanopores seem to offer more flexibility that solid state. With mutagenesis, you can get them to do almost anything.
Sat, 05 Feb 2011 20:09:23 +0000 girlscientist AGBT Mark Akeson, UCSC: strand sequencing through nanopores. I am going to defer to @nanopore for tweeting details.
Sat, 05 Feb 2011 20:08:27 +0000 rforsberg agbt Allpaths-lg needs 500 Gb memory to assemble a vertebrate genome
Sat, 05 Feb 2011 20:06:49 +0000 nanopore AGBT MA: reagents: 100ml ocean water and the air we breathe
Sat, 05 Feb 2011 20:05:22 +0000 nanopore AGBT Mark Akeson, UCSC: two types of nanopore DNA seq: exonuclease and strand. His talk about strand
Sat, 05 Feb 2011 20:05:11 +0000 apfejes http://blog.fejes.ca/?p=351 AGBT Notes on David Jaffe's talk on assembly (allpaths-lg) : http://blog.fejes.ca/?p=351
Sat, 05 Feb 2011 20:04:22 +0000 deannachurch AGBT Mark Akeson: on Nanopores!
Sat, 05 Feb 2011 20:04:01 +0000 bffo AGBT Ok boys and girls, I'm out of here! It was a fun meeting, and I enjoyed the tweets, and I like where tweeting will be next year!
Sat, 05 Feb 2011 20:02:14 +0000 deannachurch AGBT @djschlesinger de novo gets you insertions, inversions and translocations too- really is the best way to understand variation.
Sat, 05 Feb 2011 19:59:44 +0000 bffo AGBT Did Jaffe just say: if it is in genbank it must be real? :-)
Sat, 05 Feb 2011 19:59:43 +0000 djschlesinger AGBT de novo assembly identifies things (large deletions only?) not found any other way
Sat, 05 Feb 2011 19:58:33 +0000 girlscientist AGBT Take note kids, that's the pickup line for the hoe-down tonight, "don't you want to fully know my genome?"
Sat, 05 Feb 2011 19:58:08 +0000 girlscientist AGBT Jaffe, We all should be interested in denovo assembly "to fully know a person's genome."
Sat, 05 Feb 2011 19:57:11 +0000 deannachurch AGBT @djschlesinger It is the technical term!
Sat, 05 Feb 2011 19:56:47 +0000 djschlesinger AGBT Is "crappy" a quantitative term
in bioinformatics?
Sat, 05 Feb 2011 19:52:27 +0000 KamounLab http://genomebiology.com/2010/11/S1/P3 AGBT Genome Biol "Analyzing and minimizing bias in Illumina sequencing libraries"
Sat, 05 Feb 2011 19:51:22 +0000 girlscientist AGBT David Jaffe offering detailed comparisons of genome metrics by multiple assembly algorithms. Somebody's gotta do it.
Sat, 05 Feb 2011 19:50:03 +0000 genomeresearch http://bit.ly/fcj5Fd AGBT SOAPdenovo
Sat, 05 Feb 2011 19:49:38 +0000 djschlesinger AGBT Anyone have a copy of "bioinformatics for dummies" that I can borrow?
Sat, 05 Feb 2011 19:48:11 +0000 deannachurch AGBT @girlscientist yes- that is what I heard...
Sat, 05 Feb 2011 19:46:21 +0000 girlscientist AGBT Did David Jaffe just say currently ~1week for 1Gb of genome?
Sat, 05 Feb 2011 19:46:15 +0000 GenomeRef AGBT DJ: AllPathsLG- still needs significant computational power and memory: can assemble about 1G per week.
Sat, 05 Feb 2011 19:46:07 +0000 new299 http://yfrog.com/h83fapyj AGBT ALLPATHS LG running on 512Gb node for human assemblies, ouch + want!
Sat, 05 Feb 2011 19:44:28 +0000 GenomeRef AGBT DJ: Representing variation- recognizing that one path is not representative of biology.
Sat, 05 Feb 2011 19:41:32 +0000 djschlesinger AGBT Jaffe: method to increase read length is to connect 100bp paired end reads with a 3rd overlapping read from data set
Sat, 05 Feb 2011 19:37:50 +0000 deannachurch AGBT DJ: Need both technology improvements and algorithmic improvements to succeed.
Sat, 05 Feb 2011 19:37:36 +0000 djschlesinger AGBT Jaffe: de novo assembly with short reads requires new computational and new laboratory methods.
Sat, 05 Feb 2011 19:37:23 +0000 new299 http://yfrog.com/h5iqonxj AGBT ALLPATHS LG using specific lab protocol.
Sat, 05 Feb 2011 19:37:05 +0000 GenomeRef AGBT First up: David Jaffe talking about a subject we love: Genome assemblies!
Sat, 05 Feb 2011 19:36:27 +0000 girlscientist AGBT David Jaffe, Broad Inst: Goal = each new genome is not a research project. Hmmmm.
Sat, 05 Feb 2011 19:32:54 +0000 djschlesinger AGBT David Jaffe (Broad) discussing high quality draft assembly of vert genomes with NGS data
Sat, 05 Feb 2011 19:32:13 +0000 apfejes AGBT yay... more tweeting allowed for all speakers this session!
Sat, 05 Feb 2011 19:32:09 +0000 deannachurch AGBT Last session: "Genomic Frontiers" and all are tweetable!
Sat, 05 Feb 2011 19:11:12 +0000 apfejes AGBT Argh... "we searchable" should be "web searchable" notes. You'd think I'd have some practice typing by now with all that blogging.
Sat, 05 Feb 2011 19:06:47 +0000 apfejes AGBT hah. I need error correction on my tweets about error rats... or a cage to keep them in. thanks @nilshomer
Sat, 05 Feb 2011 19:04:57 +0000 apfejes AGBT @KamounLab As I said, I really enjoyed hearing it, and the blogging part just means I get we searchable notes. (-:
Sat, 05 Feb 2011 19:04:14 +0000 apfejes AGBT Any time I'm within 2m of a windows computer, the wireless becomes unreliable... coincidence?
Sat, 05 Feb 2011 18:56:04 +0000 KamounLab http://blog.fejes.ca/?p=348 AGBT @apfejes Thanks for live blogging my talk. Was fun to read your notes! http://blog.fejes.ca/?p=348
Sat, 05 Feb 2011 18:53:39 +0000 drjonboyg AGBT Off to the airport. Have safe trips home, twitterers.
Sat, 05 Feb 2011 18:39:46 +0000 omespeak AGBT? @nanopore Are you by any chance showing the system at this
Sat, 05 Feb 2011 18:39:26 +0000 halletecco agbt Thank you planning committee for picking such a phenomenal location!
Sat, 05 Feb 2011 18:20:14 +0000 neilhall_uk agbt @djschlesinger I thought I stepped into the wrong meeting?
Sat, 05 Feb 2011 18:15:07 +0000 nanopore http://bit.ly/g1HNkj AGBT If you didn't see the electronics platform that underlies Oxford Nanopore's single mol sensing: GridION
Sat, 05 Feb 2011 18:13:38 +0000 djschlesinger AGBT Must have walking into wrong room, talk seems to be about ant colonies??
Sat, 05 Feb 2011 18:11:34 +0000 djschlesinger AGBT Late to 454 workshop, don't know who's speaking.
Sat, 05 Feb 2011 17:59:41 +0000 djschlesinger AGBT Modified bases change the kinetics and the subsequent base call.
Sat, 05 Feb 2011 17:58:44 +0000 djschlesinger AGBT Steve back quickly discussing future directions in detecting modified bases.
Sat, 05 Feb 2011 17:52:25 +0000 djschlesinger AGBT Accuracy was improved to 99% with circular coverage, but read length is severely limited.
Sat, 05 Feb 2011 17:50:58 +0000 djschlesinger AGBT Gen-Probe was able to detect variants with 1% frequency with deep resequencing
Sat, 05 Feb 2011 17:49:16 +0000 djschlesinger AGBT 85% accuracy seems fixed, wonder if they've hit the threshold with single molecule sequencing technology.
Sat, 05 Feb 2011 17:43:42 +0000 djschlesinger AGBT Matt Friedenberg from Gen-Probe up next.
Sat, 05 Feb 2011 17:42:29 +0000 djschlesinger AGBT Requires micrograms of starting material. :(
Sat, 05 Feb 2011 17:42:22 +0000 apfejes AGBT did I get that right? Pac Bio has problems with GC, but was that before or after their new chemistry that fixes GC homopolymers
Sat, 05 Feb 2011 17:42:01 +0000 nanopore AGBT pm session starts 2.30, then 3pm Oxford Nanopore collaborator Mark Akeson on his work at UCSC. Focus: translocation of ssDNA thru pore
Sat, 05 Feb 2011 17:41:43 +0000 djschlesinger AGBT Bias summary: No amplification bias, but limited coverage bias.
Sat, 05 Feb 2011 17:40:31 +0000 djschlesinger AGBT coverage is consistently random across genome, but dips a bit in higher GC genomes
Sat, 05 Feb 2011 17:38:07 +0000 djschlesinger AGBT Accuracy does not decrease with read length, 85% accuracy holds out to 6kb.
Sat, 05 Feb 2011 17:36:57 +0000 djschlesinger AGBT JGI observed no GC bias out to ~85% GC
Sat, 05 Feb 2011 17:36:46 +0000 apfejes AGBT,makessense Interesting changes in Pac Bio's story this year: lower accuracy is acceptable - use platform in combination with others.
Sat, 05 Feb 2011 17:35:55 +0000 djschlesinger AGBT JGI confirms average 1.5kb read length, with longer reads out to 6kb!
Sat, 05 Feb 2011 17:33:03 +0000 djschlesinger AGBT Len Pennacchio (JGI) talking about closing gaps is microbial genome sequences with PacBio platform
Sat, 05 Feb 2011 17:27:31 +0000 djschlesinger AGBT higher error rates are tolerated better when mapping longer reads (>500kb)
Sat, 05 Feb 2011 17:25:26 +0000 djschlesinger AGBT 1Kb read with 10% error maps like a 740bp read
Sat, 05 Feb 2011 17:24:14 +0000 djschlesinger AGBT Couldn't find an alignment algorithm what could support long read lengths with indels, so the designed their own
Sat, 05 Feb 2011 17:21:30 +0000 djschlesinger AGBT Single molecule sequencing platforms dominated by insertion/deletion errors. As expected.
Sat, 05 Feb 2011 17:16:54 +0000 djschlesinger AGBT John Sorenson (PacBio) discussing informatics of their recent Cholera project
Sat, 05 Feb 2011 17:15:03 +0000 djschlesinger AGBT Commercial release expected to have 35-45Mb of mappable data
Sat, 05 Feb 2011 17:13:18 +0000 djschlesinger AGBT Will have an output of 22Mb per run
Sat, 05 Feb 2011 17:12:19 +0000 girlscientist AGBT At PacBio workshop.
Sat, 05 Feb 2011 17:11:49 +0000 djschlesinger AGBT PacBio machine will have average 1.5kb reads on commercial release, with 85% accuracy
Sat, 05 Feb 2011 17:10:02 +0000 djschlesinger AGBT 3rd gen single molecule platforms will have inherently higher error compared to 2nd gen, which looks at an average over many molecules
Sat, 05 Feb 2011 17:06:10 +0000 djschlesinger AGBT Steve Turner up discussing recent advances in PacBio technology.
Sat, 05 Feb 2011 17:02:53 +0000 djschlesinger AGBT Farewell to those Tweeps departing today. Hope you make it back safe and on time. Was a pleasure Tweeting with you.
Sat, 05 Feb 2011 16:57:19 +0000 djschlesinger AGBT Pacific Biosciences workshop about to start, hoping to see some real data!
Sat, 05 Feb 2011 16:54:48 +0000 suganthibala AGBT. Thanks to all tweeters from the forbidden zone :)
Sat, 05 Feb 2011 16:54:00 +0000 omespeak http://yfrog.com/gzyw9fej AGBT
Where the conference attendees spend time when not at a session:
Sat, 05 Feb 2011 16:51:58 +0000 omespeak AGBT Where the conference attendees spend time when not at a session:
Sat, 05 Feb 2011 16:50:41 +0000 apfejes AGBT We're slowly coming to the end of the talks, with only a few more to go this aft, but thanks to everyone for the RTs!
Sat, 05 Feb 2011 16:47:26 +0000 apfejes http://blog.fejes.ca/?p=348 AGBT Notes on AGBT talk: Sophien Kamoun on potato pathogens: http://blog.fejes.ca/?p=348
Sat, 05 Feb 2011 16:37:08 +0000 dwmohr AGBT Wondering what/who will repopulate my microbiome once I'm done my antibiotics
Sat, 05 Feb 2011 16:19:24 +0000 omespeak AGBT Interesting variety of talks this morning: from gut micobiome to potato famine pathogens.
Sat, 05 Feb 2011 16:18:14 +0000 new299 http://yfrog.com/h0qgxmjj AGBT Am I allowed to take pictures of drawings of slides?
Sat, 05 Feb 2011 16:17:14 +0000 apfejes http://blog.fejes.ca/?p=346 AGBT Notes on James Giovannoni's talk on the Tomato Genome: http://blog.fejes.ca/?p=346
Sat, 05 Feb 2011 16:09:02 +0000 drjonboyg AGBT Apparently several people saw my name in one of John Oliver's (NABsys) slides. No idea how, wonder if he'd give me a copy?
Sat, 05 Feb 2011 16:06:20 +0000 drjonboyg AGBT @nanopore well, I was at that talk so I really should have noticed it was Gundlach and not Akeson! Will follow the talk at the airport
Sat, 05 Feb 2011 16:06:06 +0000 bffo http://bit.ly/cxbIQE AGBT James Giovannoni talking about closing gaps with ngs data, referee to IMAGE software
Sat, 05 Feb 2011 16:03:45 +0000 GenomeRef AGBT JG: working to make a 'gold standard' tomato genome!
Sat, 05 Feb 2011 16:02:13 +0000 nanopore AGBT @dijonboyg @omespeak Jens Gundlach spoke Thursday (as you say MspA pores) and Mark Akeson was moved to Sat.
Sat, 05 Feb 2011 15:59:11 +0000 drjonboyg AGBT @nanopore @omespeak Akeson also spoke on Thursday, didn't he? (MspA pores).
Sat, 05 Feb 2011 15:55:42 +0000 drjonboyg AGBT I think there might be a candidate for LATFH here at (although he maybe saying the same thing about me)
Sat, 05 Feb 2011 15:55:38 +0000 GenomeRef AGBT James Giovannoni talking about sequencing the tomato genome. Started BAC by BAC, now adding NGS
Sat, 05 Feb 2011 15:54:52 +0000 omespeak AGBT After Rob Knight's talk this morning, am wondering what microbes get transferred to my fingers when using a public computer.
Sat, 05 Feb 2011 15:48:31 +0000 apfejes AGBT We've been reminded that photography of slides is not allowed - but does U2 count?
Sat, 05 Feb 2011 15:44:39 +0000 new299 http://yfrog.com/h4b7opj AGBT U2 at
Sat, 05 Feb 2011 15:38:12 +0000 drjonboyg http://j.mp/fllU7j AGBT Although actually it's not even from @nanopore ()
Sat, 05 Feb 2011 15:35:20 +0000 apfejes http://blog.fejes.ca/?p=338 AGBT For entertainment, some analytics data for my blog during season: http://blog.fejes.ca/?p=338
Sat, 05 Feb 2011 15:34:59 +0000 drjonboyg AGBT Rather bummed that the only talk I have any interest in seeing today (@nanopore) is on after I need to head to the airport.
Sat, 05 Feb 2011 15:23:23 +0000 KirstinWrites AGBT @drjonboyg I'm not at the conference ... but I agree with the statement.
Sat, 05 Feb 2011 15:11:49 +0000 omespeak AGBT Baylor U heavily invested solving the viral metageome of Elephant Herpes Virus as a step towards cure.
Sat, 05 Feb 2011 15:11:47 +0000 girlscientist AGBT Both Knight & Petrosino agreed with audience Q that w/their strategies, we'd miss viral/bacterial genomes w/weird, non-seq'able bases.
Sat, 05 Feb 2011 15:11:39 +0000 apfejes http://blog.fejes.ca/?p=335 AGBT Notes on Joseph Petrosino's talk on Viral metagenomics: http://blog.fejes.ca/?p=335
Sat, 05 Feb 2011 15:09:46 +0000 omespeak AGBT Practical application of viral metagenomics: develop therapy for Elephant Herpes virus that kills Asian baby elephants.
Sat, 05 Feb 2011 15:09:25 +0000 drjonboyg AGBT One more good use of twitter at conferences: an excellent way of meeting people.
Sat, 05 Feb 2011 15:08:42 +0000 obahcall AGBT Petrosino: Virus protocol is able to differentiate stool and nasal wash samples. We can tell our nose from our rear.
Sat, 05 Feb 2011 15:05:54 +0000 omespeak AGBT How to tell the difference between your nose & other end? Different virus exist in stool & nasal washes -J Petrosino.
Sat, 05 Feb 2011 15:05:29 +0000 new299 http://yfrog.com/h31wnpuj AGBT Nose v arse detection via microbiome sequencing
Sat, 05 Feb 2011 15:05:24 +0000 girlscientist AGBT Sorry tweeps wifi just died so radio silence ensuing.
Sat, 05 Feb 2011 15:04:34 +0000 bffo AGBT JP "pretty cool that the phage can tell you which bacteria re there"
Sat, 05 Feb 2011 15:02:44 +0000 new299 http://yfrog.com/gy5b3clj AGBT Random primers for looking at microbiome... http://yfrog.com/gy5ebblj
Sat, 05 Feb 2011 15:01:31 +0000 obahcall AGBT Petrosino: What does your poo tell you? Looking at representation of DNA/RNA virsues in stool.
Sat, 05 Feb 2011 14:58:23 +0000 obahcall AGBT Joseph Petrosino: Overview of NIH￢ﾀﾙs Human microbiome project, showing bacterial communities differ at different body sites.
Sat, 05 Feb 2011 14:57:28 +0000 bffo AGBT HMP data is so fresh, it is "dripping wet" JP
Sat, 05 Feb 2011 14:56:24 +0000 obahcall AGBT Knight: Shows a memorable visualization following the distribution of bacteria across human face.
Sat, 05 Feb 2011 14:56:13 +0000 girlscientist AGBT Joe Petrosino showing figure of vaginal viral microbiomes, with two different collection methods called "swab" & "broom." Eek.
Sat, 05 Feb 2011 14:55:29 +0000 bffo AGBT,HMP @girlscientist what about the biofilm on my iPad?
Sat, 05 Feb 2011 14:52:34 +0000 bffo http://commonfund.nih.gov/hmp/ AGBT Joe Petrosino gave intro to NIH's Human Microbiome project and also viruses ...
Sat, 05 Feb 2011 14:52:33 +0000 girlscientist AGBT [now paranoid about pushing my glasses up on my nose and introducing fingertip bacteria to site.]
Sat, 05 Feb 2011 14:48:05 +0000 girlscientist AGBT J Petrosino: human microbiome project, multiple sides probed in ￢ﾀﾜworst doctor￢ﾀﾙs appointment ever￢ﾀﾝ for subjects.
Sat, 05 Feb 2011 14:47:12 +0000 obahcall AGBT Rob Knight: Overview microbiome studies, open microbiome initiative, partnership with Gates for pers medicine in developing nations.
Sat, 05 Feb 2011 14:45:34 +0000 omespeak AGBT Joseph Petrosino: very hard to follow a talk that is probably trending on twitter.
Sat, 05 Feb 2011 14:42:55 +0000 omespeak AGBT Very entertaining talk by R Knight on microbiome diversity, especially his digression into the Onion-like NGS techniques too look for.
Sat, 05 Feb 2011 14:42:00 +0000 apfejes http://blog.fejes.ca/?p=333 AGBT Notes on Rob Knight's talk on the Human Microbiome: http://blog.fejes.ca/?p=333
Sat, 05 Feb 2011 14:41:41 +0000 new299 http://yfrog.com/h5j48qikzj AGBT Microbiome and genetics linked in fat mice
Sat, 05 Feb 2011 14:39:53 +0000 girlscientist http://www.openmicrobiome.org/. AGBT Rob Knight mentions Open Microbiome Initative,
Sat, 05 Feb 2011 14:38:18 +0000 omespeak AGBT Personalized medicine in developing nations based on microbial diversity?
Sat, 05 Feb 2011 14:33:57 +0000 girlscientist AGBT R Knight: lovely visualizations of which bacteria are on which parts of the face, but this reveals subject's habits, shall we say.
Sat, 05 Feb 2011 14:32:20 +0000 omespeak AGBT Just born babies have different microbiomes depending on normal vs c-section birth.
Sat, 05 Feb 2011 14:30:26 +0000 omespeak AGBT Wow! the biogeography of the human face is varied too, but symmetrical.
Sat, 05 Feb 2011 14:27:10 +0000 omespeak AGBT Microbiome habitats are very different in different parts of the body; also changes with time. Rob Knight
Sat, 05 Feb 2011 14:24:57 +0000 SEQanswers agbt Hopefully someone is capturing Rob Knights jokes, they are great and I can't type that fast
Sat, 05 Feb 2011 14:24:46 +0000 girlscientist AGBT Rob Knight certainly a comedian. Treating us to four proposed sequencing technologies to work out for, including unicorns.
Sat, 05 Feb 2011 14:22:51 +0000 omespeak AGBT Rob Knight: why is it called 454? It's the temp at which money burns for sequencing. :)
Sat, 05 Feb 2011 14:20:47 +0000 omespeak AGBT Micriobiomes in extreme environments are interestingly not outliers.
Sat, 05 Feb 2011 14:17:45 +0000 girlscientist AGBT Rob Knight: his keyboard microbiome study published in PNAS but then featured on CSI: Miami ￢ﾀﾜso you really know it￢ﾀﾙs true.￢ﾀﾝ
Sat, 05 Feb 2011 14:16:36 +0000 omespeak AGBT Rob Knight: each of us have an unique microbiome on fingertips that is transferred to computer keys.
Sat, 05 Feb 2011 14:13:54 +0000 girlscientist AGBT Rob Knight, UC Boulder: Problem of microbiome sequencing = get big phylogenetic trees which are hard to understand and analyze.
Sat, 05 Feb 2011 14:09:24 +0000 omespeak AGBT Rob Knight: Microbial symbionts are like unique snowflakes ie more varied than human genes.
Sat, 05 Feb 2011 14:07:35 +0000 djschlesinger AGBT Need a break, will pick up after lunch.
Sat, 05 Feb 2011 14:06:28 +0000 djschlesinger AGBT @mbcf, 8 hrs from sample to data, 5min per base, ALL reagents in single cartridge, no cBot or PE module needed, possible 510k approval
Sat, 05 Feb 2011 14:03:05 +0000 apfejes AGBT Yay! All four presenters this morning can be tweeted/blogged.
Sat, 05 Feb 2011 13:53:01 +0000 apfejes AGBT If anyone lost a pair of glasses last night, it's been turned in to reception. (but can you read this tweet?)
Sat, 05 Feb 2011 13:38:46 +0000 mbcf AGBT. Wake Up MiSeq: just a HiSeq Mini Me or something more? Blog on what you think is so clever about this box. "I want one." And ...
Sat, 05 Feb 2011 07:24:00 +0000 SOLiDSequencing http://twitpic.com/3wno0q PGM,AGBT Adam English of Baylor wins @iontorrent sequencer @ @LifeCorporation Triathlon. Congrats, Adam!
Sat, 05 Feb 2011 06:25:45 +0000 davcraig #AGBT
Sat, 05 Feb 2011 05:09:56 +0000 drjonboyg AGBT @genomicslawyer @lukejostins @mndoci in defense, it's not a default no tweet policy, its up to individual speakers.
Sat, 05 Feb 2011 04:42:14 +0000 drjonboyg AGBT OK, really going to bed now.
Sat, 05 Feb 2011 04:36:57 +0000 LIFECorporation http://twitpic.com/3wm67t AGBT Adam English of Baylor College of Medicine wins the PGM at the Life Technology Triathlon at Congrats!
Sat, 05 Feb 2011 03:34:49 +0000 girlscientist AGBT Mini tweetup with @deannachurch and @bffo in lobby bar. Stop by!
Sat, 05 Feb 2011 03:18:34 +0000 drjonboyg http://yfrog.us/5fdukz AGBT Calling it a night. Here's a video of the beach fireworks to keep you occupied.
Sat, 05 Feb 2011 01:53:55 +0000 apfejes AGBT Neat mechanism of disease in the Clinical Sequencing session, but I can't talk about it. DOH! only 56 people get to hear about it.
Sat, 05 Feb 2011 01:52:29 +0000 omespeak AGBT Last talk of the day on Cloud solutions for genomic data storage/transfer. Not doing the answer is 'cloudy since we can't tweet' jokes
Sat, 05 Feb 2011 01:52:26 +0000 dsexton_2 #agbt The Broad is testing their pipelines on Amazon. They will be optimizing Picard for the cloud.
Sat, 05 Feb 2011 01:45:45 +0000 drjonboyg AGBT There seem to be a certain irony in someone presenting on cloud computing but not allowing social media.
Sat, 05 Feb 2011 01:36:46 +0000 apfejes http://blog.fejes.ca/?p=328 AGBT Notes on Praveen Cherukuri's talk on allele specific expression http://blog.fejes.ca/?p=328
Sat, 05 Feb 2011 01:26:02 +0000 dsexton_2 #agbt everyone needs to "value add" for their cores.
Sat, 05 Feb 2011 01:24:55 +0000 SEQanswers AGBT,badbatterylifebutwhocares During this break...I must express that I have a severe case of Macbook Air envy. I'm surrounded by them.
Sat, 05 Feb 2011 01:20:08 +0000 apfejes http://blog.fejes.ca/?p=325 AGBT Notes from Kateryna Makova's excellent talk on Mitochondria heteroplasmy http://blog.fejes.ca/?p=325
Sat, 05 Feb 2011 01:18:41 +0000 dwmohr core,AGBT Why is 'core' not enough? ++
Sat, 05 Feb 2011 01:07:37 +0000 drjonboyg AGBT Is nuclear transfer from one oocyte to another even legal? Seems like the sort of thing bioconservatives freak out over.
Sat, 05 Feb 2011 01:02:46 +0000 deannachurch fail,agbt Session chair
Sat, 05 Feb 2011 00:58:28 +0000 girlscientist AGBT Marc Allard, FDA: no tweetability announced so will just say that the man clearly has difficult problems to solve. Cheers to him.
Sat, 05 Feb 2011 00:54:43 +0000 new299 http://yfrog.com/h2h0hsxj AGBT Baylor sequencers + 3 ion torrent + Pacbio.
Sat, 05 Feb 2011 00:53:25 +0000 Mikegenome agbt Did I read that right that 6 illuminas = 30 SOLiDs at Baylor? Musny
Sat, 05 Feb 2011 00:47:32 +0000 new299 http://yfrog.com/h378zrj AGBT More ion torrent error rates, lots of things less than Q17? http://yfrog.com/h56f7urj
Sat, 05 Feb 2011 00:41:31 +0000 SOLiDSequencing PGM!,AGBT In it to win it? Anyone could go home w/ the @iontorrent @LifeCorporation Triathlon NOW (5-9 pm), Suite 1106.
Sat, 05 Feb 2011 00:40:14 +0000 new299 http://yfrog.com/h4niwnij AGBT Ion torrent run reports
Sat, 05 Feb 2011 00:37:21 +0000 apfejes http://blog.fejes.ca/?p=316 AGBT Notes on Daniel Neafsey's talk on sequencing parasite genomes: http://blog.fejes.ca/?p=316
Sat, 05 Feb 2011 00:36:15 +0000 drjonboyg AGBT Horrible AV feedback in the clinical session.
Sat, 05 Feb 2011 00:33:18 +0000 djschlesinger AGBT Joe Boland from NCI is wearing a t-shirt that says "Tweet Me" in huge letters. Speaking on Ion Torrent's PGM.
Sat, 05 Feb 2011 00:32:09 +0000 SEQanswers AGBT Boland: Giant T-shirt "TWEET ME"
Sat, 05 Feb 2011 00:29:23 +0000 girlscientist AGBT Neafsey and Broad colleagues make their own whole genome bait oligos from malaria parasite DNA.
Sat, 05 Feb 2011 00:25:09 +0000 new299 http://yfrog.com/h3fq0nzj AGBT I vote that all simulated data has this banner from now on
Sat, 05 Feb 2011 00:23:11 +0000 new299 http://yfrog.com/h4vgccej AGBT Errr that looks weird, polyC artefact reads?
Sat, 05 Feb 2011 00:22:20 +0000 girlscientist AGBT Daniel Neafsey, Broad: sequencing of pathogen genomes from human clinical samples, without culturing them.
Sat, 05 Feb 2011 00:21:03 +0000 new299 http://yfrog.com/h3luugj AGBT Thinks after fiddlings as good as PCR free
Sat, 05 Feb 2011 00:20:29 +0000 apfejes AGBT Nice talk by Matthew Bainbridge, (Clinical seq session), but same (untwitterable) story told by others. Missed the twitter status too
Sat, 05 Feb 2011 00:17:03 +0000 new299 http://yfrog.com/h5sxyzj AGBT Effect of Pcr on GC bias
Sat, 05 Feb 2011 00:14:40 +0000 girlscientist AGBT The elevators made me a minute late so I missed what the lovely @deannachurch announced about tweeting policy in her clinical session.
Fri, 04 Feb 2011 23:40:10 +0000 SOLiDSequencing PGM!,AGBT It's a tight race... anyone could go home w/ the @iontorrent @LifeCorporation Triathlon 5-9 pm, Suite 1106.
Fri, 04 Feb 2011 23:15:00 +0000 drjonboyg http://yfrog.com/h8asdvzj AGBT Just watched the sun set over the ocean. Haven't been able to do that since I lived in Encinitas.
Fri, 04 Feb 2011 23:10:14 +0000 ChaunceyGrattan HiSeq2000.,agbt @Lyro20 re: David Bently & 1Tbp from How did they increase output? increased cluster density?
Fri, 04 Feb 2011 22:55:46 +0000 djschlesinger AGBT Just got to check out the MiSeq in person. The design was thoroughly thought out. An amazing instrument! Can't wait to get one!
Fri, 04 Feb 2011 22:07:28 +0000 bioinfosm AGBT Broad Institute is seemingly devoting 42 CPUs to processing data from each of its HiSeqs - heard from
Fri, 04 Feb 2011 21:37:57 +0000 Lyro20 AGBT￢ﾀﾝ #Norseqcenter ￢ﾀﾜDavid Bentley: illumina HiSeq2000 exceeding 1 TERABASE sequence/run (up from 200 Gb launch spec 12 months ago).
Fri, 04 Feb 2011 21:36:10 +0000 SOLiDSequencing http://twitpic.com/3wifv5 SOLiD,AGBT. @LifeCorporation Workflow Automation: AB Library Builder featured at
Fri, 04 Feb 2011 21:28:53 +0000 SOLiDSequencing http://t CE,sequencing,AGBT @LifeCorporation 3500 Genetic Analyzer, gold standard in capillary electrophoresis. http://twitpic.com/3widvg
Fri, 04 Feb 2011 21:26:09 +0000 SOLiDSequencing http://twitpic.com/3wid4w SOLiD,AGBT,sequencing 5500 Series Sequencer featured in @LifeCorporation Suite at
Fri, 04 Feb 2011 21:11:51 +0000 apfejes http://blog.fejes.ca/?p=314 AGBT Notes from Ion Torrent Talk http://blog.fejes.ca/?p=314 ... off to the tweetup! See you there
Fri, 04 Feb 2011 21:11:42 +0000 SOLiDSequencing http://twitpic.com/3wi93q AGBT,sequencing Introducing... the @iontorrent PGM￢ﾄﾢ sequencer, "Personal Genome Machine".
Fri, 04 Feb 2011 21:11:34 +0000 KamounLab AGBT Chad Nusbaum on Ion Torrent: very low GC bias with E. coli, accuracy uniform too, less bias than other technologies
Fri, 04 Feb 2011 21:10:04 +0000 SEQanswers AGBT @drjonboyg @apfejes IonT/Broad talk still going on.
Fri, 04 Feb 2011 21:08:29 +0000 apfejes AGBT. @drjonboyg Talk should be over in a couple minutes, and several of us will be there
Fri, 04 Feb 2011 21:06:50 +0000 drjonboyg AGBT @apfejes I think I'm the only person here.
Fri, 04 Feb 2011 21:06:40 +0000 KamounLab AGBT Chad Nusbaum on Ion Torrent applications: mutation validation, targeted resequencing, QC nextgen libraries, resequencing microbes...
Fri, 04 Feb 2011 21:00:04 +0000 djschlesinger AGBT Chad Nusbaum (Broad) about to show some new Ion Torrent data.
Fri, 04 Feb 2011 20:57:30 +0000 SOLiDSequencing http://bit.ly/triathlon-reg AGBT @LifeCorporation Triathlon REGISTRATION: BE THERE Fri. 5-9pm Suite 1106!
Fri, 04 Feb 2011 20:55:43 +0000 SOLiDSequencing AGBT Scores R tight 4 Triathlon - points R high tonight so U could come back & steal the lead!!! BE THERE: Fri. 5-9pm Suite 1106!
Fri, 04 Feb 2011 20:50:52 +0000 apfejes AGBT Tweetup @4pm in poster room by computers - I'll be 10 minutes late for Ion Torrent Talk, but hope to see everyone there.
Fri, 04 Feb 2011 20:41:03 +0000 bioitworld AGBT Moore's Law, what Law? David Bentley: illumina HiSeq2000 exceeding 1 TERABASE sequence/run (up from 200 Gb launch spec 12 months ago).
Fri, 04 Feb 2011 20:39:04 +0000 drjonboyg http://yfrog.com/h0x9cyij AGBT This sure beats being at work. Wait a minute, this *is* work! http://yfrog.com/h4udtwbj
Fri, 04 Feb 2011 20:32:03 +0000 apfejes http://blog.fejes.ca/?p=311 AGBT Notes on Tim Yu's talk on autism: http://blog.fejes.ca/?p=311
Fri, 04 Feb 2011 20:25:32 +0000 SOLiDSequencing AGBT @iontorrent Workshop NOW: Fri. 3:30-4:00 pm in ISLAND BALLROOM, Chad Nusbaum of Broad Inst.
Fri, 04 Feb 2011 20:22:57 +0000 delahar 1000g,Illumina?,AGBT By my count 90% of the sequence has been done by Is there a viable challenger?
Fri, 04 Feb 2011 20:22:01 +0000 omespeak AGBT Anyone knows if Oxford Nanopore is presenting anything at their suite?
Fri, 04 Feb 2011 20:14:35 +0000 apfejes AGBT @deannachurch Fantastic - we may all be a bit late, if talks run over.
Fri, 04 Feb 2011 20:12:18 +0000 omespeak AGBT No autism gene but genes; WGS + linkage analysis should help in identifying such heterogeneous diseases. - Tim Yu
Fri, 04 Feb 2011 20:11:43 +0000 djschlesinger AGBT @girlscientist Yes, but does that itself mean? Seems rather ambiguous. How can you do association studies without a clear phenotype?
Fri, 04 Feb 2011 20:11:34 +0000 girlscientist AGBT Are there other fields which use the term "bake-off" as much as genomics?
Fri, 04 Feb 2011 20:10:04 +0000 Awesomics http://t.co/dLotozd genome,sequencing #AGBT update from @gw_dailyscan:
Fri, 04 Feb 2011 20:09:34 +0000 SEQanswers AGBT TimYu: Two great examples of disorders presenting as autism, revealed as something else by WGS by @CompleteGenomic
Fri, 04 Feb 2011 20:08:08 +0000 girlscientist AGBT @djschlesinger he clarified that they use "the autisms" as there are many different spectra under one umbrella.
Fri, 04 Feb 2011 20:03:02 +0000 finchtalk #AGBT Tim Yu. Nice picture, each complete genomics sequence eliminates an excursion into the weeds of open source tools.
Fri, 04 Feb 2011 19:56:53 +0000 djschlesinger AGBT Tim Yu discussing genetics of Autism Spectral disorders. Maybe I missed it, but was Autism defined?
Fri, 04 Feb 2011 19:48:35 +0000 finchtalk #AGBT illumina talk. The spectrum of sequencing technology. Nice representation of throughput, turn around time, instruments. The future
Fri, 04 Feb 2011 19:47:22 +0000 apfejes http://blog.fejes.ca/?p=309 AGBT. Bloggable notes on David Bently's talk (Illumina) http://blog.fejes.ca/?p=309
Fri, 04 Feb 2011 19:46:11 +0000 Copenhagenomics AGBT Thanks to @rforsberg for the mention of our non-profit genomics conference in at - appreciate it! :-)
Fri, 04 Feb 2011 19:39:12 +0000 KamounLab espresso.,AGBT Thank you @CompleteGenomic for the Was much needed!
Fri, 04 Feb 2011 19:37:55 +0000 girlscientist AGBT @SEQanswers just barely! ;)
Fri, 04 Feb 2011 19:35:36 +0000 SEQanswers AGBT @girlscientist At least it's cheaper than 6 million dollars...
Fri, 04 Feb 2011 19:31:56 +0000 djschlesinger AGBT I want a MiSeq!!
Fri, 04 Feb 2011 19:31:27 +0000 girlscientist AGBT David Bentley of Illumina: we have made sequencing better, stronger, faster. [I paraphrase but you get the gist.]
Fri, 04 Feb 2011 19:27:57 +0000 djschlesinger AGBT David Bentley from Illumina discussing clinical sequencing. Tweeting OK sans patient data
Fri, 04 Feb 2011 19:26:51 +0000 SEQanswers AGBT @djschlesinger 12 channel pippin prep in development....!!
Fri, 04 Feb 2011 19:24:06 +0000 apfejes AGBT. @SEQanswers Ironic that the vendors have the most sane twittering policy... or perhaps not.
Fri, 04 Feb 2011 19:22:44 +0000 girlscientist AGBT MT @apfejes: Tweetup (to discuss policy): 4pm, Poster Room in the corner with the computers by the door. Hope to see you there!
Fri, 04 Feb 2011 19:22:39 +0000 djschlesinger AGBT￢ﾀﾝ Just wish it could run more than 4 samples, 12 would be nice: ￢ﾀﾜ@SEQanswers: Pippin prep 10kb automated gel isolation in 1.5h. Want.
Fri, 04 Feb 2011 19:20:43 +0000 SEQanswers AGBT Pippin prep 10kb automated gel isolation in 1.5h. Want.
Fri, 04 Feb 2011 19:17:34 +0000 djschlesinger AGBT￢ﾀﾝ Touchￃﾩ: ￢ﾀﾜ@SEQanswers: Everyone gushing onafternoon tweeting policy: It's a commercial session! ;)
Fri, 04 Feb 2011 19:16:17 +0000 SEQanswers AGBT Everyone gushing onafternoon tweeting policy: It's a commercial session! ;)
Fri, 04 Feb 2011 19:15:50 +0000 apfejes AGBT Tweetup: 4pm, Poster Room in the corner with the computers by the door. Hope to see you there!
Fri, 04 Feb 2011 19:15:16 +0000 trutane http://t.co/YEfdcKs genome,sequencing #AGBT update from @gw_dailyscan:
Fri, 04 Feb 2011 19:14:37 +0000 apfejes doingitright,AGBT Impressed with granularity in tweeting policy in afternoon sessions: don't tweet patient data when discussed.
Fri, 04 Feb 2011 19:14:12 +0000 girlscientist AGBT @apfejes tweetups seem to work best when you name a very specific place. First night's was bust bc reception was too big.
Fri, 04 Feb 2011 19:13:22 +0000 djschlesinger AGBT Chris Boles from Sage showing off the Pippin Prep for automated size selection for NGS library prep
Fri, 04 Feb 2011 19:11:04 +0000 omespeak AGBT Slightly improved tweeting policy for this session: free to blog/tweet except for where patient/clinical info is discussed.
Fri, 04 Feb 2011 19:05:56 +0000 SOLiDSequencing http://bit.ly/m-triathlon-scores AGBT @neilhall_uk You just might - you're currently in the lead: - good luck!
Fri, 04 Feb 2011 19:02:51 +0000 neilhall_uk agbt Stephan Schuster showing very nice ion torrent data....I hope I win one tonight
Fri, 04 Feb 2011 19:02:35 +0000 apfejes http://blog.fejes.ca/?p=292 AGBT Partial notes on Eric Boerwinkle's talk http://blog.fejes.ca/?p=292
Fri, 04 Feb 2011 19:00:12 +0000 apfejes AGBT Anyone intersted in tweetup @4pm, after talks? Would be nice to present unified feedback to conference organizers.
Fri, 04 Feb 2011 18:54:59 +0000 finchtalk #AGBT Steven Schuster. Helico bacter, is a genome from hell. Doing pilots on ion torrent.
Fri, 04 Feb 2011 18:53:56 +0000 SOLiDSequencing AGBT @iontorrent Workshop Fri. 3:30-4:00 pm ISLAND BALLROOM, Chad Nusbaum of Broad Inst.
Fri, 04 Feb 2011 18:49:43 +0000 SEQanswers AGBT,caffeinated @lukejostins with 12.5 hours of talks per day...they are essential
Fri, 04 Feb 2011 18:44:29 +0000 SageScience1 Sage Science Workshop 2:00pm Island Ballroom. Whats new with the Pippin Prep? Come check it out!#AGBT
Fri, 04 Feb 2011 18:42:24 +0000 lukejostins #AGBT must be pretty hectic for both @NEBiolabs and @CompleteGenomic to both decide "pick-me-ups" are the big sell
Fri, 04 Feb 2011 18:39:23 +0000 obahcall AGBT C Bustamante: Differences in ancestry switching between individuals, likely related to diffs in demographic history of MEX and ASW.
Fri, 04 Feb 2011 18:33:58 +0000 SOLiDSequencing AGBT #FF @LifeCorporation @SOLiDSequencing @iontorrent @Grand_Challenge
Fri, 04 Feb 2011 18:33:41 +0000 obahcall AGBT Carlos Bustamante: Importance studying transethnic populations, discusses sequencing of genome of 2 individs, AA and MEX ancestry
Fri, 04 Feb 2011 18:28:46 +0000 SOLiDSequencing http://bit.ly/triathlon-reg PGM,AGBT You can still enter @LifeCorporation Triathlon & play to win @iontorrent sequencer:
Fri, 04 Feb 2011 18:25:17 +0000 SOLiDSequencing http://bit.ly/agbtmobile AGBT For @LifeCorporation events, visit from your mobile device.
Fri, 04 Feb 2011 18:22:00 +0000 SEQanswers AGBT Bustamente is a great enthusiastic speaker. Think Jack Black does worldwide admixture pop gen
Fri, 04 Feb 2011 18:20:16 +0000 Mikegenome agbt Na18507 na19240 na12878 used in Bustamante analysis of novel variants
Fri, 04 Feb 2011 18:08:12 +0000 sdcrosby AGBT On behalf of those of us still in the cold, thank you so much to all you tweeters!!
Fri, 04 Feb 2011 18:06:20 +0000 finchtalk #agbt life tech lunch. Interesting how short the life cycles are for sequencing instruments.
Fri, 04 Feb 2011 17:46:42 +0000 kmjones96 #AGBT
Fri, 04 Feb 2011 17:43:38 +0000 ChaunceyGrattan agbt How are samples labeled for BioNanomatrix NanoAnalyzer analysis?
Fri, 04 Feb 2011 17:40:05 +0000 SOLiDSequencing http://bit.ly/tri-leaderboard PGM #AGBT @LifeCorporation Triathlon Scores - LEADER in contest to win @iontorrent sequencer is...
Fri, 04 Feb 2011 17:37:58 +0000 SOLiDSequencing AGBT @LifeCorporation Lunch/Workshop Now 12-2 pm PALM ROOM: Sequencing Gets Personal. 5 Amazing Speakers - don't miss it!
Fri, 04 Feb 2011 17:00:40 +0000 KamounLab http://www.jove.com/details.stp?id=1573 AGBT Primer Extension Capture (PEC) used to enrich 40K-fold heavily degraded Neanderthal mtDNA http://www.jove.com/details.stp?id=1573
Fri, 04 Feb 2011 16:57:38 +0000 girlscientist AGBT Much of Richard Green talk on Neandertals so far has been published. Still interesting but less tweetable.
Fri, 04 Feb 2011 16:56:58 +0000 drjonboyg http://j.mp/g3IhKJ AGBT This makes a good companion piece to the current talk on extinct hominid genomes:
Fri, 04 Feb 2011 16:55:01 +0000 finchtalk #AGBT RG. majority of sequences from neaderthal bone (~83%) match nothing in the DBs, others microbes, but get enough to seq the genome
Fri, 04 Feb 2011 16:53:31 +0000 owen_w agbt @deannachurch Thanks I'm sure a copacetic policy can get put together.
Fri, 04 Feb 2011 16:52:46 +0000 obahcall AGBT E Boerwinkle: Progressing from GWAS to medical resequencing to experimental systems to clinical epidemiology, with egs from ERIC.
Fri, 04 Feb 2011 16:47:45 +0000 SEQanswers AGBT Gout effect SNP rs2231142 (in urate transporter ABCG2) is in 23andMe. Just checked myself...so cool
Fri, 04 Feb 2011 16:47:12 +0000 finchtalk #AGBT Richard Green, evolution from extinct hominids. What can we learn from the dead?
Fri, 04 Feb 2011 16:45:23 +0000 djschlesinger AGBT Richard Green (UCSC) up next. No stated Tweeting policy, however from UC, so probably open to Tweeting.
Fri, 04 Feb 2011 16:43:09 +0000 genomicslawyer http://bit.ly/gFx0hR AGBT, At @GenomeQuest announces clinical diagnostic/analysis platform for whole-genomes/exomes:
Fri, 04 Feb 2011 16:42:41 +0000 KamounLab http://bit.ly/h8SyeX AGBT "Waiting for the (#Genomic) Revolution" in Science this week /via @EricTopol @GenomeScience
Fri, 04 Feb 2011 16:42:19 +0000 deannachurch AGBT @owen_w Message heard and I will bring this up with the other members of the committee.
Fri, 04 Feb 2011 16:40:07 +0000 girlscientist AGBT E Boerwinkle: "gout is the Rodney Dangerfield of diseases."
Fri, 04 Feb 2011 16:36:11 +0000 apfejes http://i.imgur.com/auLiQ.png AGBT Counterpoint, but not really differnt MT @owen_w: commentary on blogging policy.
Fri, 04 Feb 2011 16:31:14 +0000 larry_parnell AGBT Thanks to all Bloggers/Tweeps! Please keep discussing great work of those who allow & ignoring those who wish 2 stay w/in 4 walls
Fri, 04 Feb 2011 16:26:52 +0000 owen_w http://i.imgur.com/auLiQ.png #agbt My commentary on the blogging policy.
Fri, 04 Feb 2011 16:17:37 +0000 seqcentral #agbt folks: any chance of you all having an "informal formal" discussion on tweeting/blogging policy for *future* conferences?
Fri, 04 Feb 2011 16:17:15 +0000 SEQanswers http://genome.gov/GWAStudies/ AGBT Love the GWAS summarizing figure @ , but colors in legend make Tufte shiver
Fri, 04 Feb 2011 16:13:43 +0000 drjonboyg AGBT @Comprendia @mndoci it's at the speakers' discretion.
Fri, 04 Feb 2011 16:11:56 +0000 Comprendia agbt What's the latest on social media policy? @mndoci
Fri, 04 Feb 2011 16:11:06 +0000 NEBiolabs #AGBT attendees: Need a pick-me-up? Check inside your NEBNext cup for a free coupon to pick up an iced coffee or other beverage!
Fri, 04 Feb 2011 16:10:53 +0000 djschlesinger AGBT Eric Boerwinkle (UTexas) phoning in his talk and says we can "tweet him to death"
Fri, 04 Feb 2011 15:44:20 +0000 completegenomic agbt If you need a morning pick-me up, espresso is flowing at Complete Genomics suite - Everglades Room
Fri, 04 Feb 2011 15:41:56 +0000 drjonboyg AGBT @apfejes I'm happy to talk about it if you want to find me.
Fri, 04 Feb 2011 15:39:46 +0000 owen_w agbt @apfejes: "List of speakers who don't allow public discussion"
Fri, 04 Feb 2011 15:39:29 +0000 owen_w agbt @apfejes jermdemo quote: "haha great stuff - Fejes' list of untweetable talks"
Fri, 04 Feb 2011 15:35:47 +0000 illuminainfo http://yfrog.com/h4l7ncmj HiSeq,AGBT Illumina's Expert Session on
Fri, 04 Feb 2011 15:33:06 +0000 completegenomic #AGBT Genome-scale searches for novel autism genes. Dr. Tim Yu, Harvard Medical School, to present today, 2:40-3, Island Ballroom.
Fri, 04 Feb 2011 15:32:52 +0000 drjonboyg http://www.genome.gov/10002328 AGBT @apfejes that's illegal now thanks to GINA:
Fri, 04 Feb 2011 15:31:18 +0000 apfejes AGBT. @owen_w The point wasn't to embarass the speakers - most probably don't even know what it means. Those are the holes in my notes.
Fri, 04 Feb 2011 15:26:47 +0000 owen_w agbt embarrassing speakers who dont want 2b blogged is asinine - the _organizers_ failed to prep the speakers before the meeting.
Fri, 04 Feb 2011 15:26:34 +0000 omespeak AGBT Great discussion on twitter about new-born sequencing/genomics in response to last talk. Esp follow @drjonboyg & @djschlesinger
Fri, 04 Feb 2011 15:25:55 +0000 drjonboyg AGBT @apfejes good post, but in the US, newborn screening is universal and paid for by the states, not the patients.
Fri, 04 Feb 2011 15:25:44 +0000 SEQanswers AGBT @drjonboyg Sure, more phys/patient education is a requirement. Just don't like the throw-up-hands-and-hide-info strategy.
Fri, 04 Feb 2011 15:25:29 +0000 UK_Biomek fridayreads,AGBT. and my is the stream of info coming from All sounds very interesting, wish I was there.
Fri, 04 Feb 2011 15:24:44 +0000 raindancetech http://bit.ly/hzEaBg #AGBT Visit w/ RainDance in Lanai to learn more about our microdroplet-based platform & the new DeepSeq FFPE app.
Fri, 04 Feb 2011 15:22:16 +0000 drjonboyg AGBT @SEQanswers there needs to be a huge improvement in physician understanding of genomics before that happens.
Fri, 04 Feb 2011 15:21:42 +0000 jermdemo http://blog.fejes.ca/?p=234 AGBT haha great stuff - Fejes' list of untweetable talks: http://blog.fejes.ca/?p=234 (via @apfejes)
Fri, 04 Feb 2011 15:20:12 +0000 SEQanswers AGBT Not worried about incidentalome. Much like you get conf intervals/ranges on your blood tests...WGS will be presented similarly
Fri, 04 Feb 2011 15:19:58 +0000 drjonboyg http://yfrog.com/h3utsxaj AGBT Final note on NBS and genomics: not only is it a fascinating topic, it generates awesome posters
Fri, 04 Feb 2011 15:19:17 +0000 digitalbio AGBT @mndoci According to @finchtalk there were extenuating circumstances at from weather-related travel challenges
Fri, 04 Feb 2011 15:17:40 +0000 Lyro20 AGBT￢ﾀﾝ #Norseqcenter ￢ﾀﾜ@djschlesinger: Won't the "Incidentalome" prohibit WGS of newborns, or anyone, in a clinical setting?
Fri, 04 Feb 2011 15:14:34 +0000 deannachurch AGBT Martin Hibbard from GIS: No tweeting. I guess we'll just keep discussing that last talk- which was great!
Fri, 04 Feb 2011 15:14:21 +0000 drjonboyg AGBT @djschlesinger the US has 4+ million births a year, more than half under medicaid. Stratified access and disparity is a problem
Fri, 04 Feb 2011 15:13:31 +0000 apfejes http://blog.fejes.ca/?p=234 AGBT List of speakers who don't allow public discussion @ keeps growing: http://blog.fejes.ca/?p=234
Fri, 04 Feb 2011 15:13:13 +0000 omespeak AGBT Liked how Ellen W Clayton referred to Infinite Jest and GATTACA in her talk.
Fri, 04 Feb 2011 15:12:53 +0000 drjonboyg AGBT @djschlesinger right, but there's a difference between individuals and public health, and newborn screening is public health.
Fri, 04 Feb 2011 15:12:42 +0000 SEQanswers AGBT Unfortunate that ChadN had to stop the discussion re: information sharing. Was just getting interesting.
Fri, 04 Feb 2011 15:11:54 +0000 omespeak AGBT Ellen Wright Clayton's talk was informative, entertaining and thought provoking. Also thanks to her for allowing tweeting/blogging.
Fri, 04 Feb 2011 15:11:48 +0000 obahcall AGBT Clayton: Evidence and policy based use is the key to surfing the tsunami of whole genome sequencing.
Fri, 04 Feb 2011 15:11:40 +0000 drjonboyg http://j.mp/hO2BaP AGBT We raise some of these issues in this paper: (You can tell this is a policy area I work on and care about)
Fri, 04 Feb 2011 15:10:11 +0000 omespeak AGBT +then should we share the information with patients. However, we may not have a choice, considering patients may demand it.
Fri, 04 Feb 2011 15:09:06 +0000 omespeak AGBT Summary: there will be huge amount of genomic information soon, but do we know what that information means? And if we don't know that+
Fri, 04 Feb 2011 15:08:38 +0000 rhodesmd #AGBT Ellen Clayton gave a talk on ethics of medicine when we have sequence argued against giving out all info brave talk to this audience
Fri, 04 Feb 2011 15:08:36 +0000 obahcall Ellen Wright Clayton: Surfing tsunami of whole genome sequencing, need for developing a policy consensus for access and use of results.#AGBT
Fri, 04 Feb 2011 15:08:14 +0000 djschlesinger AGBT If you can't prevent parents requesting (& getting) antibiotics for their kid's viral ear infection, how will you hold back WGS data?
Fri, 04 Feb 2011 15:07:40 +0000 drjonboyg AGBT Without saying too much, I'm confident that we'll see a lot of research into all of these questions and issues quite soon.
Fri, 04 Feb 2011 15:06:33 +0000 apfejes http://blog.fejes.ca/?p=290 AGBT: Notes from Ellen Wright Clayton on dangers of human genomic information. http://blog.fejes.ca/?p=290
Fri, 04 Feb 2011 15:05:51 +0000 SEQanswers AGBT This is a fantastic talk re:"Incidentalome". Undercurrent of overwhelmed paternalistic pessimism.
Fri, 04 Feb 2011 15:05:13 +0000 omespeak AGBT We have to be rigorous about science and policy to make best use of the upcoming data deluge. Otherwise we will drown: Ellen W Clayton
Fri, 04 Feb 2011 15:04:33 +0000 drjonboyg AGBT In my experience, many in the NBS world are currently really dismissive of WGS unfortunately. we need time to change attitudes
Fri, 04 Feb 2011 15:04:24 +0000 dwmohr AGBT Meant several good arguments FOR limiting patient access to WGS
Fri, 04 Feb 2011 15:03:49 +0000 dsexton_2 #agbt how much protection will governments give?
Fri, 04 Feb 2011 15:02:49 +0000 dwmohr AGBT If tweet policy is any indication, limiting access to WGS will be a hard sell. Several good arguments against it, however
Fri, 04 Feb 2011 15:02:49 +0000 omespeak AGBT Scientific analysis of impact of genetic variation needs to proceed at fast pace for good policy - Ellen W Clayton.
Fri, 04 Feb 2011 15:01:14 +0000 drjonboyg AGBT @djschlesinger great question. I hope so, but tbh don't feel comfortable predicting. So many things have to align (tech, ELSI, budget)
Fri, 04 Feb 2011 15:00:15 +0000 djschlesinger AGBT Ellen: Physicians can't transform SNP data into clinically actionable results.
Fri, 04 Feb 2011 14:59:47 +0000 omespeak AGBT Clayton: Perhaps most controversial - since genetic information *will be* available, patient's desire for info could play minor role.
Fri, 04 Feb 2011 14:59:28 +0000 SEQanswers AGBT "What the patient WANTS will play a minor role" (in genetic information sharing)
Fri, 04 Feb 2011 14:59:03 +0000 drjonboyg AGBT One more thing: there's a difference between individuals sequencing their newborns, and population-level public health NBSeq
Fri, 04 Feb 2011 14:58:03 +0000 djschlesinger AGBT 10+ yrs away? @drjonboyg....I want to see newborn WGS happen population-wide, but there's a LOT that needs to happen first.
Fri, 04 Feb 2011 14:55:37 +0000 omespeak AGBT Clayton: Disclosure of all (genomic) information threatens to sweep away the health-care system
Fri, 04 Feb 2011 14:55:34 +0000 drjonboyg AGBT Don't get me wrong, I want to see newborn WGS happen population-wide, but there's a LOT that needs to happen first.
Fri, 04 Feb 2011 14:54:31 +0000 omespeak AGBT Clayton: Bad science is a problem. Even if we know everything about genome, we cannot predict all disease like they mention in GATTACA
Fri, 04 Feb 2011 14:54:24 +0000 drjonboyg AGBT @djschlesinger certainly something that needs to be worked out. Also, how do you get meaningful informed consent for WGS in 15 min?
Fri, 04 Feb 2011 14:53:24 +0000 djschlesinger AGBT Absolutely! @drjonboyg: Also, clinical WGS needs to get error rates down by three or more orders of magnitude for NBS
Fri, 04 Feb 2011 14:52:15 +0000 omespeak AGBT Clayton: As we look at genomic testing, vast majority of findings will have pleiotropic affects.
Fri, 04 Feb 2011 14:51:49 +0000 drjonboyg AGBT Also, clinical WGS needs to get error rates down by three or more orders of magnitude for NBS
Fri, 04 Feb 2011 14:49:55 +0000 omespeak AGBT Ellen Wright Clayton: debate on incidental findings - how much information do you give back?
Fri, 04 Feb 2011 14:49:40 +0000 drjonboyg AGBT As an example, Illinois balked at deploying a new method of NBS because it would have cost $5 million to conduct the pilot.
Fri, 04 Feb 2011 14:48:36 +0000 djschlesinger AGBT Won't the "Incidentalome" prohibit WGS of newborns, or anyone, in a clinical setting?
Fri, 04 Feb 2011 14:47:55 +0000 drjonboyg AGBT Not sure I agree that we're going to have routine newborn WGS until the costs come down to $200 or less, the States can't afford it.
Fri, 04 Feb 2011 14:46:14 +0000 SEQanswers AGBT Ellen Clayton: In a few years, newborns will be sequenced when they can't get away.
Fri, 04 Feb 2011 14:44:45 +0000 omespeak AGBT Ellen Wright trying to make a case that complete disclosure WGS data could lead to disaster.
Fri, 04 Feb 2011 14:44:37 +0000 dsexton_2 #agbt Ellen is an expert in the bioethics around sequencing
Fri, 04 Feb 2011 14:44:04 +0000 djschlesinger Ellen Wright Clayton (Vanderbilt) wants to "stir things up". Tweet away......#AGBT
Fri, 04 Feb 2011 14:35:33 +0000 KapaBiosystems AGBT Come visit our poster to see latest improvement to NGS sample prep workflow.
Fri, 04 Feb 2011 14:35:10 +0000 KapaBiosystems AGBT Library quantification kit compatible for Ion Torrent instrument. Library quant. kits available for all current NGS platforms.
Fri, 04 Feb 2011 14:25:30 +0000 neilhall_uk agbt this is all published work..why cant we tweet.?
Fri, 04 Feb 2011 14:24:29 +0000 MaliciaRogue http://lnkd.in/GXnuW3 AGBT . ￢ﾙﾻ @genomeresearch: 60 Complete, High-Coverage Human Genomes -- for Study by the Global Research Community...
Fri, 04 Feb 2011 14:10:45 +0000 wyattsgirl AGBT Following the Tweets... wish more folks could give more info to those waitlisted... ha! dang forbidders of the tweet!
Fri, 04 Feb 2011 14:09:06 +0000 deannachurch AGBT Jim Lupski up first thing this AM. No tweets b/c he is talking about patient data.
Fri, 04 Feb 2011 14:07:12 +0000 djschlesinger AGBT Dr. Lupski up first, Tweeting prohibited due to the inclusion of patient data.
Fri, 04 Feb 2011 13:46:59 +0000 BioAcousticGal slow,agbt Sounds like we missed a good one! @neilhall_uk start this morning due to the lifetech party last night...ouch!
Fri, 04 Feb 2011 13:26:29 +0000 neilhall_uk agbt #slow start this morning due to the lifetech party last night...ouch!
Fri, 04 Feb 2011 12:51:39 +0000 completegenomic #AGBT Customer case studies to be presented in Everglades Room today: 10:35-10:50, 1:30-1:45, 5-5:15. Still need an iPad? - drawing @ 6!
Fri, 04 Feb 2011 12:24:04 +0000 raindancetech AGBT Join us @ our workshop today @ 2pm to learn about Ultra-Deep Sequencing & Methylation Analysis of FFPE Tumor Samples w/ Microdroplets
Fri, 04 Feb 2011 12:19:19 +0000 illuminainfo AGBT Illumina will be at the Bronze Sponsors workshop today at 2:00 p.m. Island Ballroom
Fri, 04 Feb 2011 12:18:32 +0000 mndoci AGBT Strata videos go live a day after talks, has a default no blogging policy. Something in the world of science is just not right
Fri, 04 Feb 2011 11:42:14 +0000 pjacock http://galaxy.psu.edu/gcc2011/ AGBT Galaxy 2011 Community Conference, May 25-26, Lunteren, The Netherlands
Fri, 04 Feb 2011 06:49:48 +0000 bffo AGBT, Great PB user group evening, as well as good LT soiree sorry I missed the tweetup, maybe tomorrow evening?
Fri, 04 Feb 2011 06:02:20 +0000 SOLiDSequencing http://fb.me/Quw7A76y AGBT Stay Fresh & Focused During Yoga w/ @iontorrent every morning Capri 2 & 3, 7:00 - 8:00 AM
Fri, 04 Feb 2011 05:43:03 +0000 SOLiDSequencing AGBT Fri. @LifeCorporation Lunch/Workshop 12-2 pm PALM ROOM: Sequencing Gets Personal. 5 amazing speakers - don't miss it!
Fri, 04 Feb 2011 05:37:54 +0000 SOLiDSequencing AGBT,PGM @LifeCorporation Triathlon Scores just in! Does Baylor keep lead in competition for @iontorrent sequencer...???
Fri, 04 Feb 2011 04:44:25 +0000 abhishekpratap http://lnkd.in/GXnuW3 AGBT 60 Complete, High-Coverage Human Genomes -- for Study by the Global Research Community -- By Complete Genomics :
Fri, 04 Feb 2011 04:42:49 +0000 girlscientist AGBT, If you lose AT&T service on your iPhone here at go to settings then restore network settings. Thanks Shawn Levy for tip!
Fri, 04 Feb 2011 04:04:54 +0000 omespeak AGBT Any other tweeps at the All Night Long party?
Fri, 04 Feb 2011 02:59:08 +0000 infoecho tweetleak,AGBT It seems that we need a for
Fri, 04 Feb 2011 02:24:51 +0000 apfejes AGBT A big thank you and round of aplause for the speakers who allowed twittering/blogging today!
Fri, 04 Feb 2011 02:22:19 +0000 KamounLab http://www.bionanomatrix.com/userfiles/videos/Stretching_400kb_DNA.flv AGBT Watch bionanomatrix movie "400 kb DNA entering nanochannel"
Fri, 04 Feb 2011 02:10:34 +0000 apfejes http://blog.fejes.ca/?p=267 AGBT Notes on Maria Mendez-Lago, BC Cancer: http://blog.fejes.ca/?p=267
Fri, 04 Feb 2011 02:09:56 +0000 rafalwoycicki PAG,AGBT I have just forgotten Who can help me? What animal genome announced at was done with optical mapping by @OpGen & @BGI_events ? ?
Fri, 04 Feb 2011 02:03:59 +0000 djschlesinger AGBT Was BioNanomatrix the star this year?
Fri, 04 Feb 2011 02:03:14 +0000 KamounLab http://trinityrnaseq.sourceforge.net/ AGBT Link to Brian Haas Trinity package for RNA-Seq de novo assembly [coming soon]
Fri, 04 Feb 2011 01:48:13 +0000 drjonboyg AGBT This session has been the highlight of so far. Probably because I'm a total geek.
Fri, 04 Feb 2011 01:47:34 +0000 drjonboyg AGBT Anyone else think these nanochannels look like the matrix?
Fri, 04 Feb 2011 01:45:23 +0000 girlscientist http://www.scilifelab.uu.se/ AGBT Kerstin Lindblad-Toh asked me to pass on link to new Science for Life Lab being formed in Uppsala:
Fri, 04 Feb 2011 01:43:50 +0000 djschlesinger AGBT Han Cao from BioNanomatrix showed a cool movie of 400kb DNA fragment traveling through a nanochannel.
Fri, 04 Feb 2011 01:39:34 +0000 SOLiDSequencing http://bit.ly/lt-leaderboard AGBT.,PGM @LifeCorporation Triathlon tonight Who gets closer to winning an @iontorrent sequencer?
Fri, 04 Feb 2011 01:33:47 +0000 apfejes AGBT No blogging on Michelle Sam's talk.
Fri, 04 Feb 2011 01:30:51 +0000 apfejes http://blog.fejes.ca/?p=257 AGBT Notes on Lara Bull-Otterson's talk on finding viruses in unmapped reads for cancer. http://blog.fejes.ca/?p=257
Fri, 04 Feb 2011 01:28:59 +0000 deannachurch AGBT JO: solid state nanopore, hyb a probe and measure probe loc. //concept
Fri, 04 Feb 2011 01:28:17 +0000 girlscientist AGBT The Apple haters are winning: AT&T no service specifically for our iPhones/iPads. Can anyone explain it?
Fri, 04 Feb 2011 01:26:49 +0000 djschlesinger AGBT Solid state versus protein nanopores. which do you chose?
Fri, 04 Feb 2011 01:26:00 +0000 djschlesinger AGBT @SEQanswers likely true, but probably not enough. Point is kind of moot because the threat of bioterrorism is over stated.
Fri, 04 Feb 2011 01:24:21 +0000 SOLiDSequencing http://bit.ly/badproject AGBT Somehow escaped it thus far? Zheng Lab, Lady Gaga Parody / Science Grad School "Bad Project"
Fri, 04 Feb 2011 01:23:31 +0000 deannachurch agbt JO: Not a bioinformatics problem, not enough info content in current reads to get the right answer at all length scales.
Fri, 04 Feb 2011 01:22:32 +0000 SEQanswers AGBT @djschlesinger I agree, but you heard her mention her database...which is prob stuffed with all examples of relevant biothreat bugs
Fri, 04 Feb 2011 01:20:42 +0000 djschlesinger AGBT @SEQanswers microbial genomes are littered w/ mobile element. task is impossible, unless you link it to a modification already publish
Fri, 04 Feb 2011 01:18:58 +0000 deannachurch agbt John Oliver from NABsys on chr lenght contigs. Tweet friendly
Fri, 04 Feb 2011 01:18:05 +0000 omespeak AGBT Ironically it was a speaker from US army who was ok with tweeting/blogging.
Fri, 04 Feb 2011 01:14:12 +0000 SEQanswers AGBT,thearmyisscary @djschlesinger Probably looking for some combination of mobile elements, pieces of vectors, or novel junctions.
Fri, 04 Feb 2011 01:13:35 +0000 omespeak AGBT Lauren McNew from US military's Chemical Bio Centre - 'if you can put a sequencing machine to a camel we will buy it'
Fri, 04 Feb 2011 01:13:22 +0000 deannachurch AGBT LM: looking for a field seq unit, preferably in camo.
Fri, 04 Feb 2011 01:12:44 +0000 apfejes http://blog.fejes.ca/?p=255 AGBT Notes from Obi Grifith's talk on molecular predictors for drug response: http://blog.fejes.ca/?p=255
Fri, 04 Feb 2011 01:12:12 +0000 djschlesinger AGBT How do you tell if a microbial genome has been "engineered"?
Fri, 04 Feb 2011 01:05:35 +0000 djschlesinger AGBT Lauren McNew from ECBC can sequence a microbial genome in 36 hours!
Fri, 04 Feb 2011 01:05:18 +0000 girlscientist AGBT Obi Griffith on cancer genomics: RNAseq analysis for breast cancer subtypes.
Fri, 04 Feb 2011 01:03:16 +0000 rforsberg agbt McNew talk; finally a lab that can handle both genomics and explosives
Fri, 04 Feb 2011 01:02:25 +0000 deannachurch AGBT LM: exercises to plan for identifying pathogens in case of a biothreat, using 454.
Fri, 04 Feb 2011 01:00:27 +0000 deannachurch AGBT Laura McNew tech session. Threat analysis- cool sounding!
Fri, 04 Feb 2011 00:58:00 +0000 drjonboyg AGBT A particularly cool talk on a novel nanopore for sequencing that I can't tell you about.
Fri, 04 Feb 2011 00:38:32 +0000 djschlesinger AGBT @omespeak not so much product placement, but it's patronizing pop-culture tone. While cool, it wasn't suitable for the audience.
Fri, 04 Feb 2011 00:38:15 +0000 omespeak AGBT At the Technology session: talk about nanopores. No tweeting allowed :(
Fri, 04 Feb 2011 00:37:56 +0000 suganthibala AGBT MT @girlscientist Freudian slip? Salzberg means to say he'll give us cursory description of Bowtie, instead says "curse." You decide.
Fri, 04 Feb 2011 00:37:03 +0000 rforsberg http://cphx.org/ AGBT As of this year, Europe will also have a genomics conference inspired by AGBT. It is called Copenhagenomics -
Fri, 04 Feb 2011 00:36:22 +0000 omespeak AGBT Will talk about the movie and PacBio in details later. Some people seem pissed about the product placement.
Fri, 04 Feb 2011 00:35:47 +0000 SEQanswers AGBT Genomic Technology Session = Full. Tweets = Empty by request of speaker.
Fri, 04 Feb 2011 00:35:22 +0000 neilhall_uk agbt Well done to James hayfield for getting in a question on how to end world hunger at the pacbio dinner
Fri, 04 Feb 2011 00:34:38 +0000 omespeak agbt Couldn't Tweet from the Pac Bio dinner & movie. But movie was an interesting documentary about 'new biology' (mostly systems).
Fri, 04 Feb 2011 00:30:18 +0000 apfejes AGBT One more speaker who can't be blogged: Olivier Harismendy.
Fri, 04 Feb 2011 00:29:46 +0000 JoPet39 Not that dinner and a movie wasn't captivating, but coffee should flow freely when evening sessions are scheduled.#AGBT
Fri, 04 Feb 2011 00:29:34 +0000 SOLiDSequencing SOLiD,sequencing,AGBT Stop by Suites 1104-1106 for @LifeCorporation Triathlon and to see + @iontorrent instruments.
Fri, 04 Feb 2011 00:24:50 +0000 nanopore AGBT Jens Gundlach up now on nanopore research in the Island Ballroom
Fri, 04 Feb 2011 00:15:37 +0000 rdeborja AGBT, At the cancer genomics talks at my F-Cancer t-shirt was a huge hit. @letsfcancer, @msfuckcancer
Fri, 04 Feb 2011 00:02:06 +0000 drjonboyg AGBT Explain to me how my blackberry can have an AT&T signal (edge) when neither my iPhone or iPad have one?
Thu, 03 Feb 2011 23:43:55 +0000 SOLiDSequencing AGBT @LifeCorporation Triathlon: Stage 1 REPEAT: 7-11pm Suite 1104, Stage 2: 7-11pm Suite 1105.
Thu, 03 Feb 2011 23:40:53 +0000 SOLiDSequencing http://find.lifetechnologies.com/Triathlon-register #AGBT @LifeCorporation Triathlon, chance to win an @iontorrent PGM sequencer:
Thu, 03 Feb 2011 23:38:29 +0000 rforsberg agbt Would love to see the Monsanto single-corn-chip machine run though. Looked like something out of Brazil, the movie
Thu, 03 Feb 2011 23:36:36 +0000 drjonboyg AGBT Left the PacBio thing before the questions, had a feeling it might have gotten snarky.
Thu, 03 Feb 2011 23:36:06 +0000 rforsberg agbt It seems pretty old fashioned to use "new" biology to select solely for yield in food crops
Thu, 03 Feb 2011 23:28:42 +0000 drjonboyg AGBT Famine only happens when food shortage combines with a government preventing people from moving.
Thu, 03 Feb 2011 23:28:21 +0000 Mikegenome agbt So for a free meal we watched a 30 min PacBio commercial
Thu, 03 Feb 2011 23:27:11 +0000 illuminainfo MiSeq,illuminalounge.,AGBT Meet and greet the instrument at the Fri (2/4): 6-7pm, Sat (2/5): 1-5pm
Thu, 03 Feb 2011 23:24:49 +0000 drjonboyg AGBT Ok, that one I take issue with. Famines are political in nature, they're not going to be solved by GM crops.
Thu, 03 Feb 2011 23:24:11 +0000 djschlesinger AGBT A movie about corn eugenics? @drjonboyg: Actually, this would be a great movie to have shown my science policy class @UK_Patterson.
Thu, 03 Feb 2011 23:21:39 +0000 drjonboyg AGBT Actually, this would be a great movie to have shown my science policy class @UK_Patterson.
Thu, 03 Feb 2011 23:17:33 +0000 drjonboyg AGBT I'll have you all know I'm being incredibly restrained right now. I do like the CGI animations in this movie though.
Thu, 03 Feb 2011 23:15:57 +0000 SOLiDSequencing AGBT. @ChaunceyGrattan You're welcome! We love it, too. Lots of great info at
Thu, 03 Feb 2011 23:11:20 +0000 djschlesinger AGBT True story: @dwmohr: The new biology sounds alot like the old biology. It's the computers that can't keep up
Thu, 03 Feb 2011 23:09:21 +0000 SOLiDSequencing http://twitpic.com/3w7unm AGBT Accelerate Innovative Solutions: Mike Lelivelt at the @LifeCorporation @Grand_Challenge Workshop
Thu, 03 Feb 2011 23:09:11 +0000 dwmohr AGBT The new biology sounds alot like the old biology. It's the computers that can't keep up
Thu, 03 Feb 2011 23:05:09 +0000 ChaunceyGrattan AGBT this tag is kinda amazing... big shout out to the tweeps keeping me in the loop!
Thu, 03 Feb 2011 22:57:24 +0000 drjonboyg AGBT @dgmacarthur except most of the 'no tweeds allowed' speakers are academics, and most of the data is published or in the process.
Thu, 03 Feb 2011 22:55:30 +0000 dgmacarthur AGBT @drjonboyg Although, to be fair, has a far higher density of commercially sensitive material in presentations.
Thu, 03 Feb 2011 22:54:38 +0000 dgmacarthur AGBT @drjonboyg Sorry to see that attitude of speakers to twitter is so different from, say, BoG (where ~90% of speakers allowed tweets).
Thu, 03 Feb 2011 22:47:29 +0000 drjonboyg AGBT Not having Internet access has saved me from making a lot of sarcastic tweeds. Probably a good thing.
Thu, 03 Feb 2011 22:42:21 +0000 SOLiDSequencing http://twitpic.com/3w7mvt AGBT,PGM High jumper competes in @LifeCorporation Triathlon for chance to win an @iontorrent sequencer.
Thu, 03 Feb 2011 22:35:47 +0000 SOLiDSequencing NGS,AGBT @apfejes Targeted Reseq = 50%+ runs, then RNASeq, ChIPSeq, say JNoonan-Yale, RSteen-Harvard, JMullikin-NIH.
Thu, 03 Feb 2011 22:33:26 +0000 SOLiDSequencing http://twitpic.com/3w7kft AGBT Player competes in @LifeCorporation Triathlon for chance to win an @iontorrent sequencer.
Thu, 03 Feb 2011 22:29:32 +0000 SOLiDSequencing http://twitpic.com/3w7je8 AGBT. Maneesh Jain opens the @LifeCorporation @Grand_Challenge workshop
Thu, 03 Feb 2011 22:22:08 +0000 djschlesinger AGBT Any Tweeps at the PacBio dinner?
Thu, 03 Feb 2011 22:05:32 +0000 owen_w http://bit.ly/fAL1B5 AGBT @
Thu, 03 Feb 2011 22:03:47 +0000 girlscientist AGBT Awwww look at the kittehs.
Thu, 03 Feb 2011 22:02:38 +0000 KamounLab AGBT @girlscientist @deannachurch @apfejes Cute animal picture show continues. But wait we're not supposed to tweet about it...
Thu, 03 Feb 2011 21:59:00 +0000 NEBiolabs AGBT Check out the posters from New England Biolabs at From sample prep to bacteria making electricity:posters 7,63,92,155,184,199&206
Thu, 03 Feb 2011 21:51:15 +0000 apfejes internet,youredoingitwrong,AGBT This may be the first time in my life I'm being barred from twittering about cute kitten pictures...
Thu, 03 Feb 2011 21:50:56 +0000 deannachurch AGBT This session totally wins for cute animal pictures.
Thu, 03 Feb 2011 21:47:42 +0000 SOLiDSequencing http://bit.ly/m-triathlon-scores PGM #AGBT Triathlon Scores: It's not too late to take the lead and win an @iontorrent sequencer!
Thu, 03 Feb 2011 21:46:00 +0000 apfejes feb3,AGBT. @paniniVani I proposed we start using the hashtag. :-P
Thu, 03 Feb 2011 21:45:37 +0000 drio AGBT bowtie maps 3% > reads than bwa. What tool has the higher number of correct alignments? What alignments deliver t best SNP calls
Thu, 03 Feb 2011 21:44:57 +0000 apfejes AGBT Has Chip-Seq dissapeared from the map this year? No ChIP-Seq posters, and everyone just glosses over the epigenetics.
Thu, 03 Feb 2011 21:42:28 +0000 dwmohr #AGBT tweeting/blogging drives interest. Isn't that what we're after?
Thu, 03 Feb 2011 21:40:59 +0000 paniniVani AGBT @apfejes When did become Egypt? (via @ronald_duncan)
Thu, 03 Feb 2011 21:40:36 +0000 rafalwoycicki AGBT @omespeak sample prep is crucial
Thu, 03 Feb 2011 21:38:45 +0000 paniniVani AGBT Salzberg: adding onto TopHat- to look for fusion genes. "coming soon" (via @deannachurch)
Thu, 03 Feb 2011 21:37:12 +0000 owen_w agbt my view: when tweeting is outlawed, only outlaws will tweet.
Thu, 03 Feb 2011 21:37:11 +0000 girlscientist AGBT Tried to get my HudsonAlpha colleague Greg Barsh, to allow tweeting etc., but no success.
Thu, 03 Feb 2011 21:36:17 +0000 deannachurch AGBT Greg Barsh up next- no tweeting.
Thu, 03 Feb 2011 21:33:23 +0000 apfejes http://blog.fejes.ca/?p=241 AGBT Oops, wrong link - Notes on Steven Salzber's (VERY OPEN) talk: http://blog.fejes.ca/?p=241
Thu, 03 Feb 2011 21:31:34 +0000 deannachurch AGBT Salzberg: Getting Bowtie2 requirements during Q&A :)
Thu, 03 Feb 2011 21:30:48 +0000 djschlesinger AGBT Salzberg makes me rethink my career I should have been a bioinformatician.
Thu, 03 Feb 2011 21:27:35 +0000 omespeak AGBT Unlike, last year, no major announcements at , other than @CompleteGenomic's release of 60 human genomes.
Thu, 03 Feb 2011 21:23:09 +0000 SEQanswers AGBT Next package in the Tuxedo suite: Handcuffs. Harrrrrr.
Thu, 03 Feb 2011 21:21:20 +0000 deannachurch AGBT Salzberg: adding onto TopHat- to look for fusion genes. "coming soon"
Thu, 03 Feb 2011 21:21:12 +0000 girlscientist AGBT Salzberg: coming soon to a computer near you is TopHat-Fusion! Special-purpose routines for finding gene fusions in seq data.
Thu, 03 Feb 2011 21:21:04 +0000 neilhall_uk agbt Tophat fusion coming soon for finding fusion genes
Thu, 03 Feb 2011 21:17:37 +0000 rdeborja AGBT Bowtie2 definitely sounds interesting. Finally full support for indels.
Thu, 03 Feb 2011 21:16:54 +0000 neilhall_uk agbt Bowtie2 coming in spring
Thu, 03 Feb 2011 21:16:50 +0000 OoohMySeat AGBT. tweet censoring My view: It takes time for some to get comfortable with the power of twitter. Respect them, they will come around.
Thu, 03 Feb 2011 21:16:11 +0000 djschlesinger AGBT Bowtie 2 available this spring.
Thu, 03 Feb 2011 21:15:37 +0000 deannachurch AGBT- @rafalwoycicki not anti-tweet rules. Speaker opt-in policy. No rules broken.
Thu, 03 Feb 2011 21:15:26 +0000 djschlesinger AGBT Salzberg: Bowtie and BWA don't always map the same reads. ~7% difference.
Thu, 03 Feb 2011 21:15:05 +0000 ronald_duncan AGBT @apfejes When did become Egypt?
Thu, 03 Feb 2011 21:14:48 +0000 neilhall_uk agbt For the non-cs crowd (like me) that was a great explanation of bs transform. Thanks Steven
Thu, 03 Feb 2011 21:14:23 +0000 deannachurch AGBT @girlscientist aligns more reads- I haven't seen a measure of accuracy yet.
Thu, 03 Feb 2011 21:14:16 +0000 rafalwoycicki AGBT Thank to everyone who break the anti-tweet rules at
Thu, 03 Feb 2011 21:13:28 +0000 girlscientist AGBT Salzberg: Bowtie program still faster than SOAP2 and BWA programs for alignments, and is more accurate.
Thu, 03 Feb 2011 21:08:29 +0000 girlscientist AGBT Freudian slip? Salzberg means to say he'll give us cursory description of Bowtie, but instead says "curse." You decide.
Thu, 03 Feb 2011 21:07:03 +0000 omespeak agbt Most impressed by Life Tech's Starlight Quantum dot 'nanosequencing machines'. But have heard the talk by Beecham earlier.
Thu, 03 Feb 2011 21:05:03 +0000 Sequilabs http://bit.ly/fWov7H> AGBT Sequilabs: CompleteGenomic: New genomes will be available through our website < and t... http://bit.ly/eah5uv
Thu, 03 Feb 2011 21:05:03 +0000 Sequilabs http://bit.ly/dR4B1P AGBT, Sequilabs: CompleteGenomic: Need a caffeine pick-me up at Expresso in Everglade: CompleteGenomic: Need a ...
Thu, 03 Feb 2011 21:05:03 +0000 Sequilabs http://bit.ly/hruGBg AGBT Sequilabs: CompleteGenomic: Interested in our commercial sequencing services? Visit our hospitality suite ...
Thu, 03 Feb 2011 21:04:52 +0000 omespeak AGBT NABSys showed *some* data on nanopore sequencing by hybridization. Still no real sequencing information.
Thu, 03 Feb 2011 21:04:27 +0000 rdeborja AGBT Sitting at Steven Salzberg's High Performance Alignment and Analysis of Next Generation Sequences talk on Bowtie
Thu, 03 Feb 2011 21:04:14 +0000 deannachurch http://tinyurl.com/4lc427m AGBT Salzberg: software discussed in talk.
Thu, 03 Feb 2011 21:03:22 +0000 girlscientist AGBT Salzberg covering three of his assembly programs: Bowtie, Tophat, and Cufflinks. (what's next? Spats? Insert your own joke here.)
Thu, 03 Feb 2011 21:02:18 +0000 girlscientist AGBT Next speaker, Salzberg, says "I'd be delighted if you tweet and blog whatever you'd like about my talk,"
Thu, 03 Feb 2011 21:02:12 +0000 deannachurch AGBT Steve Salzberg: High-Performance Alignment and analysis of NGS. tweetable...
Thu, 03 Feb 2011 21:01:53 +0000 omespeak AGBT Lots of posters at focused on improved sample prep for various platforms. Very few on new sequencing techniques.
Thu, 03 Feb 2011 21:00:12 +0000 girlscientist AGBT Ssecond attempt at tweetup tonight at lobby bar, 9:30pm after night session.
Thu, 03 Feb 2011 20:58:45 +0000 omespeak AGBT General feeling from talking with various people at - sharing of and moving NGS data between various sites will be a huge issue.
Thu, 03 Feb 2011 20:54:54 +0000 apfejes http://blog.fejes.ca/?p=237 AGBT Some thoughts on talks that are forbidding tweeting/blogging. http://blog.fejes.ca/?p=237
Thu, 03 Feb 2011 20:47:48 +0000 a__muse AGBT Looks like is asking people not to tweet. So I guess blogging is also out @omespeak ?
Thu, 03 Feb 2011 20:35:29 +0000 bioscribe AGBT Dinner and a movie tonight @ featuring "The New Biology" documentary by PacBio. Palms Ballroom 5:00 pm.
Thu, 03 Feb 2011 20:22:25 +0000 KamounLab AGBT New genomics Institute at Sweden's Uppsala University modeled on the Broad. Recruiting now.
Thu, 03 Feb 2011 20:20:27 +0000 girlscientist AGBT Kerstin ends with saying new institute modeled on Broad is starting in Uppsala and they are looking for bright young scientists.
Thu, 03 Feb 2011 20:19:34 +0000 apfejes wishuwerehere,AGBT I'm sure this part is Tweetable: inbreeding dogs is bad, cancer is complex in dogs too. The rest is secret. (-:
Thu, 03 Feb 2011 20:17:12 +0000 neilhall_uk agbt It's submitted for publication but we can't tweet it!
Thu, 03 Feb 2011 20:16:11 +0000 rafalwoycicki AGBT,PAG I should be there now at the same was with
Thu, 03 Feb 2011 20:10:25 +0000 raindancetech #AGBT @ 7:30pm Thurs. - UCSD's Oliver Harismendy on analyzing chromosomal 'hot spots' in tumors w/ RainDance's DeepSeq FFPE Solution
Thu, 03 Feb 2011 20:09:00 +0000 SEQanswers AGBT I have canine compulsive disorder to tweet the details about this talk.
Thu, 03 Feb 2011 20:03:24 +0000 Sequilabs http://bit.ly/fWov7H> AGBT CompleteGenomic: New genomes will be available through our website < and the Bionimbu... http://bit.ly/ihvh0D
Thu, 03 Feb 2011 20:03:24 +0000 Sequilabs http://bit.ly/fXbuda AGBT, CompleteGenomic: Need a caffeine pick-me up at Expresso in Everglade: CompleteGenomic: Need a caffeine pi...
Thu, 03 Feb 2011 20:03:24 +0000 Sequilabs http://bit.ly/eDEdln AGBT CompleteGenomic: Interested in our commercial sequencing services? Visit our hospitality suite in the Ever...
Thu, 03 Feb 2011 20:01:24 +0000 apfejes http://blog.fejes.ca/?p=234 AGBT Not happy about it, but this is my list of talks for which I'm not allowed to publish my notes: http://blog.fejes.ca/?p=234
Thu, 03 Feb 2011 19:59:29 +0000 apfejes topsecret,canine,genome,AGBT Yet another no-tweet... this time on dog genomics.
Thu, 03 Feb 2011 19:59:12 +0000 neilhall_uk agbt. Been looking at the illumina miseq at Very very nice piece of kit. But I almost object to them making sequencing too easy.
Thu, 03 Feb 2011 19:54:57 +0000 SOLiDSequencing cancer.,AGBT New term: chromothripsis: a single catastrophic event leading to chromosomes getting interlinked in
Thu, 03 Feb 2011 19:50:30 +0000 Mikegenome agbt @drjonboyg my iPhone lost signal too. Had it in earlier sessions. Wifi working
Thu, 03 Feb 2011 19:48:42 +0000 Mikegenome agbt Blocked from tweeting what turns out midway through the Wein. talk to be published data... Perhaps the second half is unpublished.
Thu, 03 Feb 2011 19:45:33 +0000 drjonboyg AGBT, Furthering connectivity problems at now I can't get an AT&T signal on my iPad or iPhone. eek.
Thu, 03 Feb 2011 19:44:08 +0000 razZ0r AGBT Follow the hashtag for new stuff on genomes, biotech, genome sequencing, etc. (If the speaker allows tweeting, stupid policy, LOL.)
Thu, 03 Feb 2011 19:43:56 +0000 girlscientist AGBT Weinshilboum cites a recent paper, Nov 2010, in J Clin Oncol so I'd say one should look there if one wants an idea of his talk.
Thu, 03 Feb 2011 19:40:09 +0000 girlscientist AGBT Next talk, Kerstin Lindblad-Toh, not going to be tweetable either.
Thu, 03 Feb 2011 19:34:32 +0000 SEQanswers agbt First talk after poster session: Richard Weinshilboum = no tweets.
Thu, 03 Feb 2011 19:33:11 +0000 apfejes AGBT Gah... no blogging for Richard Weinshiboum. Not sure he knows what twittering is, tho... (joke about issues with the mouse...)
Thu, 03 Feb 2011 19:33:11 +0000 girlscientist AGBT First talk at afternoon session is Richard Weinshilboum, but he's requested no tweeting.
Thu, 03 Feb 2011 19:29:50 +0000 LIFECorporation http://twitpic.com/3w66nz AGBT, Getting ready for tonight's reception at hope to see you here.
Thu, 03 Feb 2011 19:29:39 +0000 cliffmcc #AGBT is asking people to not tweet during conference talks. Guess they can't take notes either?
Thu, 03 Feb 2011 19:26:18 +0000 LIFECorporation http://twitpic.com/3w65q2 AGBT Tonight's event, will we see you there? @solidsequencing @iontorrent

Not at AGBT, Day 3

2011-02-04T23:08:00.002-05:00

Okay, it's become clear that I've let the twittering controversy pretty much take over. But, why stop now? Actually, two additional points on that score (though also chuckling at the sniping at someone talking about cloud computing who wouldn't allow blogging).

First, an argument given for closing some talks is the discussion of clinical data. Some talks were shut down entirely, whereas others asked that only the clinical parts be withheld. This seems like a rather flimsy fig leaf. Any details discussed before an audience of a few hundred people can't be considered very private. As some have commented, at AGBT (and I have certainly seen this at other meetings) folks were photographing away during sessions under some degree of closure. Blogging bans primarily affect the honest and really are little protection from anything being disseminated.

Second, Owen White made a reasonable argument that any ire should be redirected from the speakers to the conference organizers. In his view, some speakers may have felt ambushed and indeed may be unaware of Twitter or how it might be something more than frivolous.

On a more positive front, one speaker actually phoned in his talk due to travel difficulties and apparently did a fine job. Another speaker (Joseph Boland) wore a t-shirt reading "Tweet me!". Curiously though, I don't believe anyone did past noting the shirt!

One thing from an earlier day (Day 1, I think) that I've been mulling is some discussion that exome sequencing seems to periodically miss regions and it isn't clear whether these are truly deletions or just sporadic dropouts. Also, full genome sequencing apparently gives more even coverage of the genome. If this is all true, then the advantages of full genome sequencing over exome sequencing are becoming more pronounced, at a time where the differential in cost is shrinking. Will targeted capture of human exomes be a phase that lasts only a year or two and is then displaced?

On the other hand, one of the talks Anthony Fejes blogged on was work at the Broad using the same capture methodology to pull parasite DNA away from their hosts, in this case Plasmodium (malaria) from human. This can be done with synthetic DNA, or by converting the genome en masse to capture probes.

Another bit of Anthony's notes I mined a bit was a talk on Ion Torrent given by Chad Nusbaum of the Broad. One interesting claim is that the sequences show very little GC bias. I'm left wondering why this plays out this way; is there a trick to their emulsion PCR? Will Life be able to boomerang that back to SOLiD? On the other hand, the quality values are apparently trending on the low side. As I've stated before, Ion Torrent (and other new platform makers) really need to push data out early & often -- they'll win more users by being upfront (we'll find the right applications!) than they'll lose. Also, they'd do themselves a favor by getting more of their customers to map their sequencers; right now there are only two locations with PGMs mapped (though AGBT could be mined to pin a few more).

Ion Torrent apparently also suggested they will launch a new chip every 6 months. If they really can get their 3rd chip out this summer, then they could pull ahead of MiSeq before any of those boxes hit the street. Of course, MiSeq will still have the advantage of the huge ecosystem of Illumina-compatible kits and protocols. Ion Torrent isn't doing themselves any favors on starting to level that playing field; if anyone can find information on how to design amplicon libraries for Ion Torrent on their website they deserve a prize. Ion Torrent also claimed they will get the 8 hour sample prep down to 2 hours; that would certainly be a big difference.

I've skipped over a bunch of other good summaries on Fejes.ca.

Yearning to Be at AGBT, Day 2

2011-02-03T22:42:00.003-05:00

Day 2 of AGBT had a Twitter feed stuffed with complaints about the Twitter/blogging policy at AGBT. It's an opt-in, not opt-out, policy. While many understood why Rick Wilson presenting a colleague's unpublished data shouldn't be tweeted out of respect, there was clearly frustration at talks which didn't have an obvious reason -- especially ones dominated by (or perhaps entirely?) published data. Plus, a number of those monitoring twitter confused the policy for a no-blogging rule. Anthony Fejes has been blogging from the meeting and is keeping a running list of who isn't allowing tweeting/blogging. Tonight's sessions seemed to be particularly untweetable.

Alas, some of the talks I wanted to sit in on weren't tweetable -- Amanda is very disappointed that the first dog talk of the conference couldn't be yipped (but is supposedly in press). But, even with the hush-hush over some talks. Apparently another talk (or the same one) full of kitten pictures couldn't be mewed either.

Another big problem at the meeting has been poor WiFi connectivity -- and some trying to use cellular data connections have had problems too. Not an uncommon problem with public WiFi, but a bit embarassing at a tech-centric meeting.

An odd Twitter comment was one claiming more tweets about the parties than the meeting itself; someone is watching a different Twitter feed than I am! On a positive note, #AGBT seems to have completely squashed #AGBT2011, other than a few holdout mugwumps -- nobody is tagging only with the longer version.

At the other end of things, rather odd for one vendor (who shall go nameless here) posting a bad photo (rotten angle, terrible lighting) of their poster via Twitter -- why not post either a good picture or better yet the PDF of the poster!!

On the positive side, Fejes.ca has nice notes on a number of talks. A few highlights extracted: Steven Salzberg has announced that the next version of Bowtie will fully support indels. He also presented data suggesting it is still much faster than some of the competing aligners (such as BWA) yet has similar or perhaps even better sensitivity. He also apparently had a very nice backgrounder on the Burrows-Wheeler transform that is the core of Bowtie, BWA and several other fast aligners -- I think I understand it but could certainly use a good tutorial.

Lots of good cancer talks it seems; Fejes.ca has notes on each. Really, really wish I were there. Also an interesting stat: Broad apparently devotes 42 CPUs to processing data from each HiSeq. YIKES!

Most of the talks tonight were apparently tweetless, but BioNanomatrix apparently impressed with a movie (unlike a much complained about one from PacBio) of 400Kb DNA moving through a channel. Not clear how close this is to practical utility.

On the vendor side, the big news is Complete Genomics releasing 40 genomes this month and 20 more next month. These include a 17 member 3-generation family from CEPH and two trios. Multiple ethnic groups are represented (Northern European, Italian, Mexican, Chinese, Japanese, Yoruban and two different tribes from Kenya). Their announcement is the first I've heard of their open source toolkit CGATools for accessing their data. Also, the data is not only on their FTP site but made available through a cloud environment called Bionimbus -- something I'll need to look at further.

One final Twitter note: I suspect that none of the vendors who hoped to have their products highlighted at academic talks are happy that most of these talks went tweetless. On the other hand, a lot of companies did not allow tweeting of their own talks, but these were early stage companies who don't yet need sales.

Oh, one tidbit from yesterday I forgot to mention. One group (I think it was Sanger) was doing something with BACs for physical mapping. The one tweet I saw wasn't detailed (are they ever?) -- was this with arrayed BACs or in solution pools that have never been picked? Would be curious to know for something else I'm writing for this space.

One final note -- tried out my Perl code and discovered there is a 100 tweet return limit. So, modified the code to step through to older tweets. Here is the new version which will slurp up to 10K messages (just modify the loop variable max to go even crazier; AGBT is only around 500 right now).
#!/usr/bin/perl
use Net::Twitter::Lite;

my $nt = Net::Twitter::Lite->new(
username => $ENV{'TWITTER_USERNAME'},
password => $ENV{'TWITTER_PASSWORD'}
);

my $queryTerm="#AGBT -RT";
my $searchHash={'q'=>$queryTerm,'rpp'=>200};

my $outCount=0;
my $lineCount=0;
my @statuses=();
for (my $i=0; $i<10; $i++)
{
my $r=$nt->search($searchHash);
last if (scalar(@{$r->{'results'}})==0);
push(@statuses,@{$r->{'results'}});
print "#\t",$i,"\t",$statuses[$#statuses]->{'id'},"\n";
#exit;
$searchHash->{'max_id'}=$statuses[$#statuses]->{'id'}-1;
}

foreach my $status(@statuses)
{
my $origText=$status->{'text'};
$lineCount++;
next if ($origText=~/^RT/);
my $text=$origText;
my $url="";
if ($text=~m/(http:[^ ]+)/)
{
$url=$1;
$text=~s/ ?$url//;
}
my @hashes="";
while ($text=~m/ \#([^ ]+)/g)
{
push(@hashes,$1);
}
$text=~s/\ #([^ ]+)//g; $text=~s/ +$//;
print join("\t",$status->{'created_at'},$status->{'from_user'},$url,join(',',grep(/[a-z]/i,@hashes)),
$text),"\n";
$outCount++;
}
print STDERR "Done! $outCount/$lineCount output\n";

AGBT From Afar, Day 1

2011-02-02T23:19:00.002-05:00

Well, I'm not at AGBT. The sobering fact is that even if I had registered, I wouldn't be at AGBT -- Logan Airport closed yesterday morning and won't open until sometime tomorrow. Even with a plow doing the driveway, there was plenty of digging to be done here.

There wasn't a lot going on with Twitter today about the conference, but a few nuggets of interest. Actually, the first order of business seemed to be a debate on whether to use the hashtag #agbt or #agbt2011. While the shorter version seems to dominate, a few folks are using the #agbt2011 alone and then there are mugwumps tweeting both. I haven't started using my Perl client yet, but this is one more advantage to having it -- I can easily merge both streams (which I can probably do with the standard interface, but sometimes it's easier to write one's own code than read someone else's documentation!)

A lot of the news out of AGBT today came from vendors -- various new announcements. For example, RainDance announced a new product for targeting FFPE DNA and is partnered with NuGen for upstream DNA amplification. LifeTechnologies is giving away another Ion Torrent PGM sequencer with a conference "Triathalon" -- though my non-attendance is irrelevant since it's open only to academics (ahem!). LifeTech also announced another "Grand Challenge" contest, $1M to someone who can sequence an entire human genome AND transcriptome from a single cancer cell using SOLiD.

Talks tonight were from some of the major sequencing centers. Harold Swerdlow from the Sanger apparently covered both their current Illumina operations as well as trials of the Pacific Biosciences instrument. The Sanger is now generating a jaw-dropping megabase of data per second; when I was a grad student one of the grant proposals was to build a megabase per day facility. Or, if you'd prefer, apparently that works out to two terabases per week. Yow!

On the Pacific Biosciences side, they are apparently getting 28 million mappable bases from reads averaging 1650 nucleotides, implying around 17K reads per 40 minute run. Definitely a lot of things one could do with that, but also it won't really compete in a lot of arenas -- particularly with only 86% accuracy. Still, long noisy reads could be very useful for scaffolding smaller ones and with 500 base fragments this would mean three passes in circular consensus mode.

A few other things that caught my eye. Harvard Biopolymers uses Twitter to let people know of spare Illumina lanes. Galaxy seems to be quite a hit and has a thriving ecosystem of add ons -- and easy install on the Amazon cloud. The Broad showed even more interesting cancer data -- though no details tweeted. Actually, their multiple myeloma sequencing paper has been allegedly in press since a conference I attended last June -- will it ever see daylight?

Tomorrow should be meatier -- talks all day. Since I won't be there, I also won't be faced with the horrid dilemma posed by the concurrent sessions in the evening. One is on all sorts of blue sky sequencing technologies and might make the pulse race. The other has lots of talks on using existing technologies, first with a string of cancer talks and then a bunch of RNA-Seq. Friday has a similar split -- one session on techniques and one on applications (including 1 of two dog talks in the confernece). Still, I'd love to struggling to decide!

Thanks to everyone who has tweeted! If only there was a way to get the slides too. Some meeting organizers might worry that too much off-site access to a meeting might hurt business. But with Marco Island, it's hard to see much worry. First, it's a cool locale. Second, watching remotely isn't the same as being there. Third, we can't see or touch any hardware (both a PGM and a MiSeq are apparently onsite). Finally, those of us remoting in can't network very effectively -- and that is the most fun of such conferences.

Three Visions of The Next Phase of Sequencing

2011-01-31T23:41:00.002-05:00

When thinking about the current crop of sequencing instruments, it occurred to me that they can be divided roughly into three categories, representing three different visions of what sequencing will look like for the next few years.

In the first category you have the monster instruments. I will use as my exemplar Pacific Biosciences, though this category would also include Helicos and to some degree the mainline sequencers of the other families. These instruments are large and require their own floor space (or effectively so). Think PDP-11 as an analogy.

In the second category are the "personal sequencers". The archetype is the Ion Torrent Personal Genome Machine (PGM), but with similar form factors are the 454 Jr and MiSeq. These instruments sit atop a bit of bench space. The PGM in particular is meant to be seen: a stylish logo composed of four symbols and even an iPod dock (what next? Personalized "skins" for your PGM?). Think personal computer as an analogy (PGM is a Mac & MiSeq a PC?).

Luke Jostins has a very nice article looking at what Oxford Nanopore has released about their instrumentation. Now, the big catch is that any working chemistry to go with that instrument is still tightly under wraps (and someone from OxNano smothered speculation that any big announcement will come at Marco Island this week). What is striking about their instrument is that while it could theoretically sit on a benchtop, what they are really aiming for are huge gangs of them -- the boxes fit in a standard computer rack. Talk about a data center!

This isn't just a mounting decision; OxNano is thinking in terms of the unique properties nanopores would bring. For example, there wouldn't be set cycles -- so each unit can read from a sample until some specified condition is met (read depth, found a given mutation, etc). The units are also designed to talk to each other, enabling large jobs to truly be spread over multiple units (SeqHadoop? The OxNano Collective -- your sequences will be assimilated?).

Fundamentally, this is a very different vision of the future of sequencing than PGM. Very few biological instruments I've seen are designed to be packed en masse efficiently; Codon had stacks and stacks of off-the-shelf thermocyclers but their loading designs insisted on a racks that wasted lots of space. Such packing implies a large, centralized sequencing center. Each unit can take 96 samples, much as the PacBio can (and presumably OxNano will conveniently take 96-well plates, though in that form factor it isn't obvious how). Contrast this with the PGM, which requires each chip to be manually locked into the instrument. Perhaps PGM will later get a Big Brother, but for now it is clearly aimed at sequencing as a cottage industry.

Which gets back to an ongoing debate in science -- should science be done on a huge scale or by small teams. Should sequencers be present on every bench like a microcentrifuge or concentrated in big centers like a supercollider (or even like the autoclaves in an academic complex)? If sequencing is to become a routine and integral part of healthcare, will it be performed in doctors' offices or central labs? These will all be ongoing questions.

Of course, none of these reach the form factor we all dream of -- something the size & convenience of a tricorder.

AGBT Around the Corner

2011-01-27T00:02:00.003-05:00

In less than a week, the AGBT meeting (perhaps better known as Marco Island) begins. Alas, due to my usual dithering around going to meetings I'm not registered -- and I've been told it sold out the day registration opened. Barring some passes showing up on StubHub, I'll be stuck monitoring Twitter again.

AGBT can be a locale for making big announcements -- last year the meeting finished with Jonathon Rothberg unveiling (well, sort of) the Ion Torrent PGM and the year before PacBio was the star. But, I'm not expecting a lot of big announcements from the major players. The recent J.P. Morgan conference was the scene of Illumina's big MiSeq announcement and Ion Torrent's launch of their second generation chip. It's hard to see either of those groups making further announcements. Nor does either have a big talk to launch things, though I'm sure many vendors will be making announcements even without a podium spot. There is one talk featuring Ion Torrent from one of their early access sites, and I'm sure that will be popular for trying to get some ideas as to what really working with the instrument is like.

As far as the other established players, Roche/454 desperately needs to generate some excitement with some advancement; the long-heralded 1Kb reads might do. PacBio has two talks, one on the cholera work and one on detecting modified bases. Perhaps some more details on performance will leak out, but neither sounds promising. Complete Genomics has a workshop but no apparent talk.

The biggest opportunity for metaphorical fireworks (to go with the ones on the beach one night -- I love fireworks; absolutely must register next year!) would be if one of the nanopore or other blue sky technologies actually demonstrated real data. I'd say any read 10 bases or longer would be something, and demonstrating a set of reads in the 20+ basepair range would mean they are really on to something. There's a handful of talks scheduled for such technologies (Oxford Nanopore's founder, Nabsys, UC Santa Cruz on nanopores, BioNanomatrix; apologies if I missed someone). No talk from GnuBio; would be nice to see whether they are at all on their aggressive timeline.

I will be watching Twitter, so if you are there and can tweet it will be appreciated. I did try to follow ASHG in November on Twitter and got a bit frustrated with the standard web interface. So, today I whipped up a quick Perl harness to search for tweets and process them into a table. Most of the heavy lifting, of course, was done by a CPAN library & so I can actually put the code below in case anyone else finds it useful. Once the meeting starts, I might hack some more on it. My dream app would know the conference schedule and assign each tweet to a talk, but that's way beyond what I see myself getting together. Perhaps I'll actually go so far as to generate a SQLite database from the tweets of interest, but most likely it will just feed Excel.

Currently what it does is pull things like hashtags and URLs out of the text & put them in separate columns. The first column is the timestamp and the second is the author. Also, it skips over explicit re-tweets. One issue I may try to deal with is posts which are really re-tweets but not tagged correctly; if those are a nuisance I may try to work on some auto text-clustering. And perhaps I won't resist at least a little crude assignment of tweets to talks (looking for keywords; tracking a given user).


#!/usr/bin/perl
use Net::Twitter::Lite;

my $nt = Net::Twitter::Lite->new(
    username => $ENV{'TWITTER_USERNAME'},
    password => $ENV{'TWITTER_PASSWORD'}
    );

my $r=$nt->search($ARGV[0]);
my $nt = Net::Twitter::Lite->new(
    username => $ENV{'TWITTER_USERNAME'},
    password => $ENV{'TWITTER_PASSWORD'}
    );

my $r=$nt->search("Ion Torrent");

foreach my $status(@{$r->{'results'}})
{
    my $origText=$status->{'text'};
    next if ($origText=~/^RT/);
    my $text=$origText;
    my $url="";
    if ($text=~m/(http:[^ ]+)/)
    {
        $url=$1;
        $text=~s/ ?$url//;
    }
    my @hashes="";
    while ($text=~m/ \#([^ ]+)/g)
    {
        push(@hashes,$1);
    }
    $text=~s/\ #([^ ]+)//g; $text=~s/ +$//;
    print join("\t",$status->{'created_at'},
                    $status->{'from_user'},$url,
                    join(',',grep(/[a-z]/i,@hashes)),
                    $text),"\n";
}

Yet Another Sequencing Service Provider

2011-01-24T23:16:00.002-05:00

GenomeWeb and Matthew Herper both covered today's announcement by Perkin Elmer that they are launching a very sophisticated second generation sequencing service. Perkin Elmer becomes a rare large, publically-traded corporation in the market, though Beckman Coulter and Illumina are significant players as well (and in their specialized manner, Complete Genomics too).

Around a year ago I felt I had spoken to every U.S. based commercial provider of second generation sequencing, as well as pretty much every Boston or Cambridge core lab offering services and a few others (such as my alma mater). I've long since given up trying to maintain that status. Barriers to entry in the field seem to be minuscule, leading to new entrants on a regular basis. Many can be found through the dedicated thread at SeqAnswers or the Second Generation Sequencer Map, but just Googling for "sequencing service provider" tends to get a few new hits. Spread your scope to Europe or the rest of the world and the number of providers climbs further.

Given such a crowded field, what I would recommend to providers is they start thinking about differentiating themselves. How? Well, look for what the other providers don't offer.

For example, nobody is yet offering a low-minimum cost rapid turnaround service. Now, partly this is because platforms to support such have only recently become available. However, the one platform that is somewhat amenable to this (454 Jr) has only one service provider I've heard of, and they are offering 4 week turnaround times -- not what I call fast. For fast, I'm thinking more on the order of a week or less. On cost, there really need to be some offerings in the sub $5K range. Yes, you can buy individual lanes from some labs (particularly core facilities) for just over $1K each, but you'll need to wait until someone else fills a plate. There is a real need for some offerings using Ion Torrent, PacBio and/or MiSeq in this sort of role.

Another potential area of differentiation is sample prep. Most service providers I've talked to still want 5-10 micrograms of input DNA for their sequencing. Want to stand out? Start specializing in 50-100 nanogram input amounts. Or formalin fixed paraffin embedded samples, which most clinical folks need to deal with. There aren't many providers using the fancier PCR-based targeting schemes (Fluidigm and Raindance) nor many I know of offer automated size selection.

It is also interesting to see that new providers tend to be either purely Illumina or offering Illumina plus some other platforms. Indeed, some of the providers who previously were strong SOLiD or 454 shops seem to all be getting into the Illumina side. Another sign that Illumina is leading the pack; let's hope that continues to be good for the industry.

If you want to hear more of my thoughts on this matter, I'll be giving a talk on this topic during the last day of Bio-IT World (indeed, in the penultimate slot of the Next Generation Sequencing Informatics track).

Time to Recognize A New Norm in Scientific Review!

2011-01-23T21:45:00.004-05:00

The 20 January issue of Nature has a news article "Trial by Twitter" exploring the issue of how scientists should deal with comments on their papers on the Internet. The article covers a lot of ground but I'd like to deal with a little of it. Also, it is a news article and tries hard to be neutral; I will not be as neutral.

Part of the article touches on two life sciences papers from last year which attracted significant controversy: a genome-wide association study (GWAS) on human longevity and the claim of arsenic-incorporating bacteria. In both cases, the authors initially declined to respond to pointed critiques in blogs. I won't mince words where I stand on this: claiming it is "premature for us to talk about our experience because this is still an ongoing issue" (the GWAS authors) or "any discourse will have to be peer reviewed in the same manner as our paper was" (the arsenic authors) is stonewalling against the interest of science and should be unacceptable in the community of scientists. It is particularly ill advised to make such a statement after, as the arsenic bacterium authors did, engaging explicit courting of the popular press to publicize the work as significant and well-executed.

That said, one cannot expect scientists to know of every tweet or reference to their paper. Nor should scientists have an obligation to address comments which have no basis in science nor engage persons engaged in personal attacks. Obviously that leaves a lot of discretion -- but discretion is different than saying "all roads go through further formal peer review". It is also unacceptable to hide behind the fact that the original paper passed peer review; as a number of papers have demonstrated, peer review often fails for various reasons. The Reactome paper is a particularly painful example of this for me, since I swallowed it whole and later realized that I (and apparently the chosen reviewers) lacked the expertise to spot glaring issues in the paper. But, I do believe that scientists do have an obligation to respond to detailed, reasoned, scientific critiques of their work.

Who should decide which critiques to respond to? The article talks a bit about post-review scientific scoring systems such as the Faculty of 1000. These are useful, but not necessarily the best way to find the most energetic and informed reviewers for a paper. Among those bloggers who comment on a paper, you are likely to find such individuals, though not with a perfect signal-to-noise ratio. Given that traditional journals are constantly struggling to find ways to stay relevant in an Internet world, to me it is the editors at the journal a paper was published in are obvious candidates for selecting those critiques demanding response. Of course, the authors of the papers themselves should also play this role.

The article discusses how poor the response has been to commenting facilities at journal websites which enable such commentary. Now, my one interaction with such a site did lead to a peer-reviewed version of my blog post. But, that is the only time I've actually tried to contact the editors of a paper, and only because I felt so strongly. Editors and authors need to be more proactive, with the expected norm being that they actively look for intelligent commentary on papers they edited or authored. Resources such as Research Blogging can help find these; indeed, I would argue that any editor who doesn't scan Research Blogging for coverage of papers in their journal should immediately start doing so.

The point of scientific publishing and peer review is not to protect reputations and not to promote orthodoxy; the point of these is to attempt to ensure that good science is made better and bad science is swept clean as soon as possible. The existing system of formal review represents an approach to this goal which evolved over time; it is neither perfect nor unquestionable. The time is now for a new ethos in science in which any reasoned source of scientific criticism is accepted and can expect response.

The Emperor of All Maladies

2011-01-20T08:36:00.001-05:00

I finished yesterday Siddhartha Mukherjee's The Emperor of All Maladies. With the exception of one shocking lapse, it's a very good history of cancer therapy and I strongly recommend it for anyone interested in cancer.

Mukherjee is an oncologist and interleaves his historical view of cancer with the experiences of selected patients he tended. This humanizes the subject and brings useful context. On the historical side, he reaches all the way back to the first written description of cancer, from Pharonic Egypt (which was also the first expression of futility at treating cancer). A major thread running through the early part of the book is the career of Sidney Farber, who pioneered chemotherapy, and Mary Lasker, who reshaped the role of the U.S. government in cancer research despite having never held elective office.

The book is written for a lay audience and I think he does a good job. There are some key lessons which need to be heard widely. Mukherjee describes many of the twists and turns, the clever leaps of logic and the leaps that failed. For example, Farber's key insight was that leukemia was characterized by improperly functioning bone marrow, a trait shared with several nutritional deficiencies which had just been solved. So he tried treating childhood leukemias (deemed utterly untreatable by oncologists of the time) with B-vitamins, to disastrous results. But, then the second leap occurred -- Farber tried B-vitamin antagonists, and soon found success.

Mukherjee also captures many of the missed opportunities and the non-scientific barriers to progress. Farber was shunned by many colleagues because he was not an oncologist, but a pathologist. The initial misstep led to utter non-cooperation from the oncologists, leaving Farber's doctors to sharpen their own needles and dump their patient's bedpans. His supply of anti-folates came from an immigrant doctor who had left for chemistry and industry after his foreign credentials proved useless. Much more recently, there is the story of how Weinberg's group discovered Her-2 but didn't contemplate pursuing therapy against it. Far worse is how the lawyers at the institute Farber founded nearly iced Gleevec, and even after that issue was dealt with (when Gleevec's proponent Brian Druker moved to another institution) the management of Novartis nearly killed it. Similarly, Genentech all but dropped Herceptin; again it was an outside oncologist who drove the project to success.

The book also gives a good overview of how many things needed to be invented along the way and issues which arise. Simple epidemiology noted the high incidence of scrotal cancer in boy chimney sweeps; the intersection of a rare cancer and rare occupation made the relationship unquestionable. But later, researchers were faced with the challenge of proving a common cancer (lung) was linked to a common environmental factor (smoking) and this required new methods such as the case-control study. In the chemotherapeutic arena, the book covers many iterations of trial design and testing strategy and also brushes a bit on the changing ethical landscape.

If you've read my previous book reviews, a typical exercise for me is to ask what else could have gone in. Now, this is admittedly sometimes unfair to the author and one prior reviewee has politely chided me that some of what I missed ended up on the editor's floor. Plus, books that want to be read and not doorstops need to respect certain length limits. But, when I am writing I find it a useful exercise; only when we consider the whole range of possibilities can we be confident that the correct balance has been reached.

Perhaps the most surprising area barely touched on is angiogenesis and anti-angiogenic therapy. Some of this may have to do with timing; most of the book concerns events before about 2004, though with an impressive quick coverage of some of the learnings from cancer genomes. So many of the travails of Avastin would be after that rough divide. Still, this is a hot topic and perhaps that is why I miss it from the book. This book deserves to be widely read and probably will be (it is certainly the de facto Book-of-the-Month at work) and will therefore form the foundation for many public conversations about cancer

Another topic essentially absent from the book are various types of immunotherapy. Again, this is an area having some resurgence (with the approval of the prostate cancer vaccine Provenge), but it isn't clear how important it will be long term. I think there was a passing mention of Coley's toxins, but saw none of the late 70's excitement around interferons (which, alas, became only a niche player in oncology) or the 80's focus on interleukins.

Finally, though there is some coverage of the world of molecular analysis of cancers (such as cancer genomics), the field of microarray classification of cancer and guiding therapy isn't explored. This is a pretty complex topic with a lot of shifting (and, as noted in the recent George Poste Nature opinion piece, far more smoke than light) so it's a bit understandable it was left out.

Overall though, I can't think of anything that is clearly missing. But now my quibble and serious complaint. Certainly the book not only filled in a lot of areas I just hadn't been exposed to, but even had me running for articles very close to where I see my core training.

The quibble is a bit of a pet peeve. The book, like many (and myself), roughly divides cancer chemotherapeutics into two bins (with room for a small "unclassified" bin as well). Cytotoxics are broad-spectrum cell killers and constitute the bulk of cancer therapy agents.

Targeted therapies are the sticking point. For me, it is important to reserve this category for agents that have two critical properties: we know what the drug acts on in cells and we know something about the relevance of that mechanism to a tumor. Mukherjee gives a number of interesting stories about targeted therapies. For example, anti-estrogen therapy for breast cancer can trace back to a doctor hearing from Scottish shepherds that removing a ewe's ovaries would cause their udders to shrink. Another fascinating story relayed in the book shows how these categories can be tricky. By undescribed means (probably random screening), it had been found that cis-retinoic acid could be used to treat the aggressive leukemia APL, though with highly variable results. An inspired leap of logic led to the decision to test all-trans retinoic acid (ATRA) in APL, with stunningly successful results. Only later was it discovered that most APL cases are driven by a fusion protein derived from a retinoic acid receptor. So ATRA started as "other" and only later fits my definition of targeted therapy.

So what's my beef? It's when Mukherjee describes Velcade, thalidomide and Revlimid in multiple myeloma as targeted therapies. He's hardly alone; this is a common description. But in my taxonomy, Velcade is a cytotoxic. We know where it acts in the cell but not why that is important in cancer and not how to select which patients to use it in. Thalidomide and Revlimid probably should go into the "other" bucket; we're not sure of the mechanism but they aren't generically cytotoxic. Even one of the drugs I currently work on (an HSP90 inhibitor) I would generally call a cytotoxic; it's not a perjorative in my book. On the other hand, in one subset of lung cancer there is a strong hypothesis as to how such HSP90 inhibition works at a molecular level. So as with many biological classifications, they're a bit smudgy -- but I still think they are useful and worth being precise about.

Finally, the big complaint. One of the longer stories in the book concerns the apogee of intensive chemotherapy, in which breast cancer patients were given doses high enough to utterly destroy their bone marrow, followed by bone marrow transplants. This was an important and controversial approach to therapy, with patients begging to get into trials and fighting legal battles to have their insurance companies pay for these unproven treatments. Indeed, Masschusetts was one state which enacted legislation to mandate coverage of this particular treatment. However, when the clinical trial results rolled in, all but one large study showed no benefit. The outlier study showed a huge benefit. However, on closer inspection the outlier turned out to a fraud of truly monstrous proportions. In discussing that denouement, Mukerjee points out that a male patient "obviously" couldn't have been a legitimate member of the trials. Breast cancer in men is rare, but rare is not impossible and it is particularly critical for oncologists to not have that blind spot. Indeed, this is particularly important in families with a history of breast cancer; the risk of male breast cancer is much higher in BRCA1/2 familes.

That one blemish aside, though, I can strongly recommend this book. As I've noted before, if many read it then there will be a much stronger general basis for discussing cancer and the public policy around it.

Whither Ion Torrent?

2011-01-13T23:09:00.003-05:00

In a comment on yesterday's piece, Matthew Herper asks how the Ion Torrent might be better than Illumina's MiSeq. That's a head-scratcher, and I'm sure the Ion Torrent folks are scrambling to figure out what to tell people.

The problem is, looking at it honestly, is that Illumina is claiming they'll beat or meet Ion Torrent on every dimension. Similar startup cost (once you thrown in necessary gadgets for the Ion Torrent), similar run cost, similar raw read lengths (but paired!), similar numbers of reads, double the total output, faster runs and less hands on time or trouble. Did I leave anything out?

So where does Ion Torrent go to keep from being stillborn? Now, they say they have 60 instruments sold, which isn't quite stillborn -- but will MiSeq choke off further growth?

The one potential gap is that Ion Torrent is (almost) here now whereas MiSeq won't show up for about 6 months. That's not a big window, but some folks will be anxious enough that they'll want their sequencer NOW. But, how many sequencers can they really churn out without quality going haywire? The last thing Ion Torrent needs to put itself behind a reputational eight ball. Of course, pushing hard to place sequencers could pay off huge if Illumina is delayed in rolling out MiSeq -- or can't meet demand.

Upfront cost similarity is for one sequencer. For a variety of reasons, some folks will want more than one (e.g. to make sure at least one is already operating). Presumably the $50K increment starts applying here, though it's not clear how many Ion Torrent's can be supported by that $50K-$75K in auxiliary gear. And, it's sobering that you don't beat MiSeq on throughput until you buy the third one. Indeed, for similar amounts you could buy 5 Ion Torrents or 2 MiSeq and have similar throughput.

Giving away sequencers would be another way to grab an edge -- or cut the price even lower. Obviously, they could start with complimentary placements with a few key genomics bloggers (grin). Seriously, this is not an unprecedented model. Kodak took the world by storm giving away cameras -- processing & reload on the original Brownie was doable only at Kodak. This would be an expensive strategy, but would presumably tie down a bunch of users from going with the competition.

How quickly can Ion Torrent up their specs? That would be the best way to win -- up Illumina's ante in a huge way. This is what their contests are around, but they need the boost now. Again, it is sobering what a challenge has been put before them. Illumina has roughly 2X the read length, though as paired ends rather than one read -- but for many applications paired ends are equivalent or superior (I have one tiny one where you need a monolithic read, but I don't see it as a "killer app"). Longer reads, requiring more cycles, would also exacerbate the speed issue -- putting even more pressure on shortening cycle times.

The sample prep issue is potentially a tough nut. Illumina's slurp of Epicentre means quick-and-easy fragment library preps. For PCR amplicons, any platform will do. But, emulsion PCR seems to be something nobody loves. I haven't seen it done & I think some folks see the trouble as overblown -- but have you ever heard someone say "I love the smell of emulsion breaking in the morning! It smells of victory!"? Illumina's bridge PCR approach certainly looks easy. Can Ion Torrent get access to some non-emulsion clonal amplification (rolling circle?) and get it in place quickly enough?

The ultimate leap past Illumina would be to roll out a new chip beyond the 2nd generation one Ion Torrent just announced. Presumably Illumina has MiSeq's specs near the limits of what the great masses of users will be able to routinely get, but perhaps they left some slack in -- but 5X slack? If Ion Torrent could launch another 10X in number of sensors -- or quickly roll out a way to get more of those sensors working -- that could keep people excited.

Ion Torrent might also try some sort of ante upping in integrating with another player or a novel sample prep approach. Nothing leaps to mind as really fitting the bill, but perhaps something is out there. Hard to beat low input fragment libraries with just a bunch of pipetting, but who knows?

Finally, there is pure cool factor. Ion Torrent had it, iPod dock and all. Apple makes a mint selling cool stuff (with very slick user interfaces and integration) at a premium price. I doubt this will work in the scientific arena, but who knows?

Yep, it's quite a rabbit which Jay Flatley pulled out of his hat this week. Knocking your opponent off-balance before they actually rolled out is something executives must dream of. I think Ion Torrent will survive (and hope it will; Illumina continuing to expand its 70% market share really wouldn't be good for the market), but their trajectory is looking a lot flatter at the moment.

My, Oh My, Oh MiSeq!!

2011-01-12T00:25:00.003-05:00

I was supposed to be going off to sleep after posting my latest on Ion Torrent but made the mistake of checking my RSS feed -- and there was a new item from Forbes' Matthew Herper (who, I was thrilled to find mentioning this space last week) about a new system from Illumina (announced this evening, hence I get to scoop GenomeWeb!). So much for a quick exit to bed (luckily, I'm working home tomorrow -- Boston area expecting a vicious winter storm).

The new system is called MiSeq and it is clearly a direct challenge to Ion Torrent as well as the 454 Jr. According to the press release, it will be priced around $125K with run costs in the $400-$750 range. What's really stunning are the specs: 6.8M 2x150 paired end reads in 27 hours -- including cluster generation!! That's really astounding! Plus it needs only 2 square feet of bench space for the entire setup.

Illumina's chemistry is well established, and (as I really appreciate having just done a SOLiD project) they really have the bulk of the academic informatics mind share as well. Paired end reads are really valuable in a number of contexts, or with short targets (e.g. PCR amplicons) they can be treated close to a single read of somewhere less than 300bp.

The time is also amazing compared to HiSeq, which is spec-ed at 8 days for 2x150 paired end reads. Now, some (all?) of this speedup could simply be through shortening the imaging time; since the flowcells for MiSeq are much smaller, the time to scan them could be similarly reduced. I don't know how much time per base cycle is in scanning vs. chemistry, but it seems like a lot of time could be saved. Alas, if this is the answer (as opposed to clever speeding up of the chemistry), then it doesn't translate to faster run times for HiSeq.

The only catch is the system is being announced now; actual machines won't ship until summer. My rough memory is that Illumina has been pretty good about hitting their product launch targets, so perhaps by end of summer I could try one out. It also certainly puts greater impetus on Ion Torrent to drive performance of their system. Illumina is claiming that sample-to-data time will be faster and about 2X the data of the new chip, plus paired end information. Ion Torrent's challenge areas could tip things back the other way -- halve the sample-to-data time, double the read length and double the accuracy.

Illumina also announced they are buying sample prep kit company Epicentre. Epicentre's amazingly clever transposon-based library prep system was one ingredient in a recent whole-genome haplotyping paper (which I was supposed to write on by now; gotta get that done!). They've also demonstrated the ability of this approach to build libraries from amazingly small input amounts. It's a natural buy for Illumina (Epicentre also makes kits for mRNA and miRNA preparation), but it's also a bit bittersweet to see it happen as this means these technologies won't be available (or at least won't be developed as energetically) for other platforms. Of course, if newer platforms can use libraries designed for Illumina, that issue won't apply (as I've noted before).

Ion Torrent Throws Another Chip Into the Pot

2011-01-11T23:48:00.004-05:00

At the J.P. Morgan conference today, Ion Torrent announced the launch of their second generation chip ("316"), which offers 10X the data generation at apparently 2X the list price (some of this is based on earlier information, which isn't always consistent). Actual chips in the hands of customers is stated to occur in this quarter.

Delivering upgrades to the systems via the consumables rather than new instruments is a big part of the promise of Ion Torrent's system, and so actually delivering the chips is a key part of fulfilling that promise. The GenomeWeb article made no mention of any 3rd generation chip, and certainly it will be the regular release of upgraded chips that will really convince the community that this is for real.

The new chip is described as delivering 100Mb per run, or about 1 million reads of 100 bases each (again, ballpark). It's useful to put that in the context of various possible uses. In my eye, the Ion Torrent still will just not have the number of reads for applications requiring counting tags -- e.g. RNA-Seq or ChIP-Seq. But, for PCR amplicons this would be pretty amazing. Imagine a pool of 100 amplicons; this would mean on average 10K reads per amplicon, which would allow great sensitivity for rare variant detection (either in pooled samples or heterogeneous cancer biopsies). As far as a human genome goes, 100Mb is about 0.025X, so it would not be cost effective (vs. Illumina or SOLiD) or pleasant (imagine snapping all those chips in to the instrument!) to go for 40X human coverage. On the other hand, that is enough to give copy number profiling information. It is also about 2X a human exome chip, which isn't nearly enough either. But, for a small genome it's pretty decent -- certainly good enough for 40+X coverage of many microbes. A de novo project might need other data, but for resequencing of industrial or clinical variants this could be quite interesting.

1M reads is also in the range of what 454 Senior delivers per run. Of course, those are much longer reads -- if you have and need the length you'll care. But given the upfront cost is so much smaller & the per run cost a tiny fraction, it would suggest that the 454 platform is going to quickly be delegated to niche status. Ion Torrent has claimed in interviews that much longer reads have been seen in house, but it isn't clear when these protocols will be rolled out or if they are really robust.

The GenomeWeb item also suggests a reasonably healthy initial uptake of the platform, with 60 orders booked. However, only "early access" customers have gotten any. It is also rumored that Life Tech is strongly encouraging multiple purchases by customers; someone I know with experience in this noted that this was the pattern with their capillary sequencers. On the one hand, this has practical advantages (beyond getting a few more sales), as the rollout can be staged to a smaller number of sites initially. New technologies always have their hiccups. But, on the negative side it does mean that fewer sites will have an opportunity to put the machine through new paces -- and compete in Ion Torrent's Grand Challenges (must write more on that another time).

Ion Torrents haven't started showing up on the wonderful World Sequencer Map yet, but it should be a matter of time. More seriously, no service provider has yet announced support for this platform. That's a pity, since that would enable much wider access to the capabilities. I can't promise I'll be first in line with an order, but I certainly will be if I have a project then that fits the specs for the machine.

Chromothripsis: Cratering of Chromosomes

2011-01-10T22:48:00.005-05:00

A stark frame from Apollo 16 shows a lunar surface remodeled by violent collisions. Even in a single static snapshot hints at the order of events. The large crater near the center of the image was later remodeled by a not small crater breaking the original rim. Careful study of other photographs, especially of the even more chaotic far side of the moon, can piece together the temporal order of events, from such overlaps in craters and their ejecta.

The moon has more than a few parallels to the genomic chaos present in cancer. Most cancers are characterized by a degree of aneuploidy, though a few only have small numbers of genomic alterations, much like the smoother regions pictured. Some cancer cells have chromosome complements like the far side of the moon, battered to the brink of non-recognition. And like the two large craters, we often picture the chromosome alterations as being sequential, with long gaps in between. Such step-wise changes are seen as mirrored in stepwise changes in biology; each new change has the potential to bring new advantages to a proto-tumor cell. A corollary of this is that these changes are ongoing; today's tumor has more changes than several months ago, and a sample several months in the future will look yet different. Indeed, one criticism sometimes launched at cancer genomics efforts is that they are taking a single snapshot of a fast-moving phenomenon.

If you look closer at the lunar photo, another interesting feature will emerge. What might first appear to be a slash pointing at the large crater is actually a chain of craters. These are believed to have arisen from a single object fragmenting before impact, much as the comet Shoemaker-Levy did a number of years ago prior to crashing into Jupiter. Such crater chains are seen many times on our own moon, as well as on Mars and moons of Jupiter. A new paper in Cell brings our attention to the same phenomenon in a subset of cancers, in which genomes are multiply mangled in a single event. And, like Apollo 16, the key evidence for understanding very dynamic events are static genomic snapshots.

For the impatient such as this author, Figure 1 C&D is perhaps the quickest way to dispel doubt that such occurs. These use Circos diagrams to depict the chromomal rearrangements present in a chronic lymphocytic leukemia (CLL) sample at initial presentation and relapse: the rearrangement diagrams look like carbon copies, despite 42 distinct alterations in the diagnosis samples. These were identified by high-throughput paired-end sequencing. Many of these breakpoints are locally clustered on the original chromosome; the long arm of chromosome 4 suffered nine breaks, each re-connected to a different chromosome. Clustered breaks are sometimes remarkably tightly packed: seven rearrangments are from a single 30 kilobase region and another 6 from a 25 kilobase region. However, fragments from such near origins are flung around, often ending up distant from each other in the tumor chromosome.

Digging deeper, a curious pattern emerges from looking at the pattern of loss of heterozygosity versus copy number. Clearly the single copy regions cannot be heterozygous, but what is striking is that all of regions of copy number two are heterozygous. This clearly indicates that none of the copy number two regions are the result of a reduction to single copy followed by an expansion back to two copies; heterozygosity lost cannot reappear. Change points for copy number fall essentially evenly into the four possible intrachromosomal buckets: deletions, head-to-head inverted, tail-to-tail inverted, and tandem duplications.

Now, this could be a single odd case. The researchers combed through the masses of publically available cancer cell line copy number and LOH data (which they largely generated) to find more. Out of 746 cancer cell lines, 18 (2.4%) showed the pattern of frequent changes in copy number in local regions of chromosomes. So the phenomenon appear to not be unique to their original sample, though present in a small minority of cell lines. They selected four cell lines for paired end sequencing to further explore this.

The sequencing reveals the same pattern, though sometimes even more intensely. One line has 239 rearrangements of chromosome 15; another 77 alterations of the short arm of chromosome 9. A third has only 55 rearrangements in chromosome 5. And while these changes are not the only rearrangements in these genomes, the other changes tend to stand aloof from the chromosomes which have been extensively fractured.

Such a chromosome catastrophe (for which the authors coin the term "chromothripsis", "thripsis" is apparently Greek for shattering into pieces) could have a number of beneficial effects for a tumor, as the authors explore. One route is through the amplification of beneficial oncogenes, perhaps through the formation of small auxillary chromosomes. In a small cell lung cancer line, such a chromosome carries only 1.1Mb of original cellular content , but contains the potent oncogene MYC -- and multiple copies even.

Conversely, such a radical remodeling of a chromosome could potentially destroy multiple tumor suppressors in a single event. Possible examples of such simultaneous multiple disruptions were found in one chordoma and several other samples.

The extensive reordering of chromosomes during chromothripsis can also lead to the formation of fusion genes. But, as noted in the paper, it would seem unlikely that these are frequently tumor drivers (though never confuse unlikely with impossible!). In chronically chromosomally unstable cells, there are presumably many different possible rearrangements generated, increasing the odds of finding one with useful advantages. But with chromothripsis, the cell would have to get lucky on one mighty roll of the dice.

What could perform such a genetic wrecking act? One possibility raised in the paper, which was also the first to jump into my mind, would be high-energy ionizing radiation (probably most commonly X-rays from terrestrial sources, but also perhaps cosmic rays). Imagine a condensed chromosome and then picture an energetic particle hitting that chromosome. Some paths through the chromosome would result in one or a few breaks -- but some paths might enfilade the DNA, riddling it with breaks. Alternatively, broken telomeres can lead to the inappropriate fusing of chromosome ends, followed by rebreakage when the chromosomes are pulled to opposite poles during cell division (which in turn now have sticky ends). While this wouldn't technically be simultaneous destruction, a few successive rounds of such "breakage-fusion-bridge cycles" during cell division can generate quite a mess. It is striking and consistent with this that several examples found in the paper involve rearrangements near the end of one chromosomal arm. It will be fascinating to see if anyone tries to replicate these phenomena in controlled settings and try to create chromothripsis in the lab, and thereby determine which mechanism is more consistent with the data -- or perhaps to what fraction each mechanism occurs and any distinguishing genomic artifacts for each mechanism.

This paper has certainly altered my perspective of the chromosomal chaos seen in cancer. My previous gradualist mental model now has an asterisk. Where I previously pictured cells in a constant process of continuing rearrangement and amplification, now I can see that in some cases the genome is actually relatively static. There is a minority school of thought which has argued that aneuploidy is the critical feature of cancer and that any chemotherapeutic regimine is doomed by the dynamics of aneuploidy. Here are clearly some counterexamples; tumors which are not constitutively unstable and which do not progress after treatment due to further rearrangement. It also is a useful paper to recall the next time someone dismisses cancer genomics with the argument that cancers are dynamic, but a genome analysis looks only at one point in time. Sometimes, if you look carefully enough, a single snapshot can tell a very dynamic story.

P.J. Stephens, C.D. Greenstein, B. Fu, F. Yang, G.R. Bignell, L.J. Mudis, E.D. Pleasance, K.W. Lau, D. Beare, L.A. Stebbings, S. McLaren, M-L Lin, D.J. McBride, I. Varela, S. Nik-Zainal, C. Leroy, M. Jia, A. Menzies, A.P. Butler, J.W. Teague, M.A. Quail, J. Burton, H. Swerdlow, N.P. Carter, L.A. Morsberger, C. Iacobuzio-Donahue, G.A. Follows, A.R. Green, A.M. Flanagan, M.R. Stratton, P.A. Futreal, & P.J. Campbell (2011). Massive Genomic Rearrangement Acquired in a Single Catastrophic Event during Cancer Development Cell, 144 (1), 27-40 : 10.1016/j.cell.2010.11.055

[2011-02-03 Fixed URL for paper; Research Blogging apparently has been failing to build it correctly]

Oncogenesis Via Altered Enzyme Specificity, Part II

2011-01-06T19:51:00.003-05:00

As promised in the EZH2 story, there is another story of cancer-causing mutations tuning an enzyme in an interesting way. It's also a great story of how multiple high-throughput methods can create and exploit an entirely new angle on cancer. I'll try to do a good job on this, but I'm lucky enough to have as regular readers of this space several of the authors who are referenced here, which should enable any egregious errors on my part to be flagged. I'm also trying to tell the main thread of the story as I can see it, and apologize in advance for getting priorities of discovery incorrect. I'm relying on final publication dates for organizing the timeline, which is certainly not a perfect strategy.

First, we have to go back just over two years ago to the late summer and early fall of 2008. Two different groups reported on initial cancer genomics investigations of glioblastoma, a devasting type of brain tumor. Both groups used PCR to amplify targets for Sanger sequencing. One group looked at a focused set of genes in 91 tumor samples; the other group looked at many fewer samples (22) but at most known protein-coding exons.

Now this is the sort of decision which was critical: given a particular sequencing budget, do you sequence a lot of targets in a few patients or a select set in more patients? Given we know a lot of oncogenes and tumor suppressors, there is a logic to the focused search. But this was a case where the broad sweep paid off.

What the broad sweep found, but was not included in the focused search, were mutations in the gene for IDH1, a key enzyme in the citric acid cycle. A rapid follow-up study confirmed the recurrent presence of IDH1 mutations in glioblastoma and also mutations in IDH2, another gene encoding a homologous enzyme.

IDH1 and IDH2 encode isocitrate dehydrogenase, a key enzyme in the citric acid cycle, which is also known as the Krebs cycle or tricarboxylic acid cycle (TCA). This set of metabolic reactions is often shown near the center of a large metabolic diagram as a big circle, which is very appropriate. This set of reactions is central to aerobic energy generation as well as the creation of various useful metabolic intermediates. The normal activity of IDH is

Isocitrate + NADP+ <=> 2-Oxoglutarate + CO2 + NADPH + H+

As with all reactions of the TCA, this reaction is reversible; under some conditions some cells will convert 2-oxoglutarate to isocitrate. Also keep in mind that a common synonym for 2-oxoglurate is alpha-ketoglutarate or aKG.

Now the influence of primary metabolism on cancer is a hot topic -- again. Back in the 1920s Otto Warburg earned a Nobel prize for the observation that tumors seem to rely on anerobic glycolysis more than their normal cousins. The field was pretty cold for a long while but lately it has gotten new interest, including a number of startup companies trying to develop cancer therapeutics. During my last job interruption, I consulted for one of these (Agios), though I have no ongoing financial interest in the company.

A striking observation is the pattern of the mutations: in each case a single arginine residue is mutated, though to multiple possible amino acids. However, these are not equiprobable. In the current COSMIC, there are 1859 reported IDH1 mutations -- and 1468 (78.97%) of those are R132H. Only a handful of mutations have been found outside R132. IDH2 has two hotspot sites, R140 and R172

Now, what are these mutations doing? What is special about IDH? The first attempt to answer this was published in April 2009 and came to the conclusion that the mutant enzyme is less effective at binding its substrate and generating the product alpha ketoglutarate. Furthermore, the mutant enzyme was proposed to poison the wild type copy by the formation of inactive heterodimers. Finally, this was proposed to activate the important HIF1 transcription factor, which regulates a number of tumor-promoting pathways. So in this view of the world, IDH1/2 are tumor suppressors inactivated in glioblastoma.

The part that was unsatisfying about this explanation is that it failed to explain why IDH1/2 mutations are so focused. In general, many mutations can destroy enzymatic function, so tumor suppressor enzymes generally show a diffuse mutation pattern. It is dangerous to think we can think through such biochemical puzzles, but it did mean the solution to the puzzle wasn't a clear winner.

A very different explanation was provided by the group from Agios, published at the end of 2009. Using high-throughput metabolite profiling, their startling discovery is that the IDH1 mutations result in higher levels of 2-hydroxyglutarate (2HG), a compound structurally-related to the normal IDH product alpha-ketoglutarate.They confirmed that the mutant enzyme is no longer capable of driving the normal reaction, but that it now catalyzes an analogue of the reverse reaction which uses the aKG and NADPH generated by the wild-type enzyme to generate 2HG. Heterodimers appeared to be capable of both reactions, raising the possibility that heterodimers enable very efficient production of 2HG through the coupling of the two enzymatic activities. Structural studies supported this explanation, and finally increased 2HG levels could be detected in glioblastoma samples mutant for IDH1, but not those wild-type for IDH1.

Around the same time, another key thread entered the story. Several attempts to identify IDH mutations in other cancers had been made, and while a few had been found there wasn't an obvious cancer with a high frequency of mutations. But, the second acute myelogenous leukemia complete genome sequenced by the Wash U group identified an IDH1 mutation and went on to confirm recurrence of IDH1 mutations in just under 10% of AML samples assayed. Now a second tumor type showed IDH recurrence. Further studies identified IDH2 mutations as well in this disease and confirmed that IDH-mutant leukemias accumulate 2HG.

So now we have an odd mutation pulling and interesting trick of changing the reaction specificity of a metabolic enzyme and showing up repeatedly in two very different cancers. But why is this odd metabolite valuable to the cancer? That is where the latest paper comes in. Published last month, it demonstrated a number of features. First, leukemias mutant for IDH1 or IDH2 show a distinctive DNA methylation profile, one which is not specific for which enzyme is mutated. This methylation profile also shows a greater degree of methylation than most other AML samples. Second, the RNA expression profiles for these tumors is not quite as highly clustered. Third, expression of mutant IDH enzymes in cell lines raises the amount of 5-methylcytosine in their DNA.

The big clue uncovered is that IDH1/2 mutations are not only mutually exclusive, they are also strictly exclusive with another recurrent mutation in AML, those inactivating the enzyme TET2. More strikingly, TET2's enzymatic role appears to be the first step in demethylating DNA -- and TET2 requires alpha ketoglutarate! Indeed, co-expression of TET2 and IDH1 mutant (R132H) reduced the degree of formation of the TET2 product (5-hydroxy-methylC) vs. TET2 + wild-type IDH1. Furthermore, TET2-mutant leukemias actually show a similar methylation profile as IDH1/2-mutant leukemias.

How does this drive leukemogenesis? Looking at the differentially-methylated sites in IDH1/2 mutant AMLs versus other AMLs, an enrichment for motifs associated with the transcription factors GATA1/2 and EVI1, both known to be important in myeloid differentiation. 40% of the genes in the IDH1/2 signature are known targets of GATA2 and 19% direct targets of GATA1. Furthermore, GATA1 was hypermethylated in their patient cohort, suggesting two levels of suppression of this pathway. Finally, mutant IDH expression or loss of TET2 function was shown to generate more cells with stem-like characteristics, a hallmark of leukemias. In particular the oncogenic kinase c-KIT showed higher expression; mutational activation of c-KIT characterizes yet another subset of AML.

So in just over two years, we've gone from high-throughput sequencing finding a curious recurrent mutation, to a novel oncogenic modification of metabolism and now a mechanistic explanation of how this drives leukemias. I've left out a lot of other literature using these mutations to guide better prognosis in cancers and the identification of recurrence of IDH mutations in some other tumor types, notably thyroid tumors. Curiously, another set of thyroid tumors appear to be wild-type for IDH1/2 (at least in the hotspot) but have elevated levels of 2HG. Germline IDH2 mutations have also been identified in a subset of patients with abnormal levels of 2HG. Some patients have inactivating mutations in a different gene, succinic semialdehyde dehydrogenase; will this show up as mutant in yet another set of cancers?

So what next? Ideally the clinical value of these findings would go beyond simply staging patients. There are hints that some chemotherapies may perform better or worse in the context of these mutations. Ideally, therapies directed at inhibiting the mutant IDH activity (whilst sparing the wild-type activity) will be developed. The higher expression of c-KIT in IDH1/2 and TET2 mutant AMLs may suggest the use of c-KIT inhibitors. Certainly one suggestion is to look in other IDH1/2 mutant tumors and in 2HG-elevated IDH1/2 wild-type tumors for distinctive hypermethylation. With larger and larger mutational datasets, more mutations may be found which are clearly mutually exclusive with IDH mutations (exclusion with NPM has also been observed in leukemia); such findings could lead to identifying further genes affecting genome methylation.

Figueroa ME, Abdel-Wahab O, Lu C, Ward PS, Patel J, Shih A, Li Y, Bhagwat N, Vasanthakumar A, Fernandez HF, Tallman MS, Sun Z, Wolniak K, Peeters JK, Liu W, Choe SE, Fantin VR, Paietta E, Löwenberg B, Licht JD, Godley LA, Delwel R, Valk PJ, Thompson CB, Levine RL, & Melnick A (2010). Leukemic IDH1 and IDH2 mutations result in a hypermethylation phenotype, disrupt TET2 function, and impair hematopoietic differentiation. Cancer cell, 18 (6), 553-67 PMID: 21130701

Oncogenesis Via Altered Enzyme Specificity, Part I

2011-01-04T22:55:00.002-05:00

(Correction: a friend close to the story pointed out EZH2 is a lysine, not arginine methyltransferase. Stupid mistake! -- though I got it right once in the original version -- small consolation)

There's a bit of an involved story I've been meaning to put together & now another paper with a similar theme showed up. After some thought, I realized that the second story should go first.

Oncogenes are genes which when added to a cell can transform it to a cancerous state. A number of different classes of proteins can be oncogenic, but quite a few are either transcription factors or enzymes. I'm going to focus here on enzumes.

Oncogenic enzymes somehow have an enzymatic activity which promotes cell growth. A lot of oncogenic enzymes are protein kinases, and these can be activated by a number of mechanisms. For example, some are activating simply by being overexpressed, which in cancer occurs most commonly by amplification of the underlying chromosomal DNA. Another recurrent mechanism is the removal of inhibitory domains. Other changes alter the equilibrium between active and inactive states. Certain kinases are activated by dimerization, so some oncogenic mutations enhance dimerization. For example, in some fusion kinases, in which a chromosomal rearrangement has fused a kinase with another protein, a key role of the partner protein is to supply a dimerization motif.

The RAS family of GTPases are an interesting variant on this theme. RAS proteins (KRAS, HRAS and NRAS being the most important oncogenes) transmit growth-promoting signals when they have a bound GTP. They also have a slow GTPase activity which hydrolyzes the GTP to GDP, and when RAS proteins have GDP bound they no longer transmit the signal. Exchange of the GDP to GTP reactivates the growth signal. Oncogenic KRAS mutations slow or eliminate the GTPase activity; without this activity the gene never turns off. Hence, only a small number of possible mutations in KRAS will successfully turn it into an oncogene, since mutations must inactivate the GTPase without altering the other functions of the protein.

The two stories, one brand new related here and one which hit a fascinating milestone recently which will be in a future installment, are cases of additional ways enzymes can be altered to promote tumors. In each case, rather than activating or inactivating an enzyme the mutations succeed in tuning the activity of an enzyme in a way favorable to cancer.

About a year ago the Vancouver cancer genomics group published the identification of recurrent mutations in lymphomas of the gene EZH2, a histone methyltransferase. Strikingly, the mutations are strongly concentrated on a single change, modifying Tyr641, though to a number of other amino acids. So what is so important about Tyr641? A new paper provides the mechanistic explanation.

Histone methyltransferases such as EZH2 add a methyl group to lysine residues (other methyltransferases can methylate arginine, which I mistaken pegged EZH2 in the original version of this). Any given lysine can actually have 4 different methylation states: none, single, double, or tri. This means in turn that an lysine methyltransferase has three types of substrates: those with 0, 1 or 2 existing methyl groups. What the new work shows is that Tyr641 is important in selecting the substrates, and these mutations focus the activity on converting dimethyl lysine to trimethyl lysine.

Several lines of evidence point to this conclusion. Two other lysine methyltransferases have been shown to prefer trimethylation when mutated in an analogous way. Molecular modeling suggests that this tyrosine serves to inhibit effective operation on dimethyl substrates. In vivo the mutation acts dominantly to increase trimethylated lysine levels on histones and in vitro the appropriate complex has an increased preference for dimethylated peptides.

This is the first such reported disease-causing mutation of this sort, though as noted above similar mutations have been created by scanning mutagenesis. Will we see other ones? There are many other homologous methyltransferases, but a quick sampling of COSMIC doesn't reveal a homolog with any recurrent pattern of mutation. It's worth keeping a lookout for one, but if not then a new mystery will remain to be explored: why is EZH2 special in this regard?

Going a bit farther afield, could there be oncogenic mutations in kinases which alter the substrate specificity? Given that some kinases require prior ("priming") phosphorylation of substrates, could a mutation in the kinase reduce this requirement? Alternatively, do some kinases phosphorylate both cancer-promoting and cancer-retarding substrates? If so, could mutations exist which shift the balance towards cancer promotion? Seems like a long shot, but who would have guessed in advance of mutations like the EZH2 ones?

Yap DB, Chu J, Berg T, Schapira M, Cheng SW, Moradian A, Morin RD, Mungall AJ, Meissner B, Boyle M, Marquez VE, Marra MA, Gascoyne RD, Humphries RK, Arrowsmith CH, Morin GB, & Aparicio SA (2010). Somatic mutations at EZH2 Y641 act dominantly through a mechanism of selectively altered PRC2 catalytic activity, to increase H3K27 trimethylation. Blood PMID: 21190999

Semianalogy to the Semiconductor Industry?

2011-01-03T23:19:00.001-05:00

One area where Jonathon Rothberg has gotten a lot of mileage in the tech press is with his claim that Ion Torrent can successfully leverage the entire semiconductor industry to drive the platform into the stratosphere. Since the semiconductor industry keeps building denser and denser chips, Ion Torrent will be able to get denser and denser sensors, leading to cheaper and cheaper sequencing. It's an appealing concept, but does it have warts?

The most obvious difference is that your run-of-the-mill semiconductor operates in a very different environment, a quite dry one. Ion Torrent's chips must operate in an aqueous environment, which presumably means more than a few changes from the standard design. Can any chip foundry in the world actually make the chips? That's the claim Ion Torrent likes to make, but given the additional processing steps that must be required it would seem some skepticism isn't out the question.

But perhaps more importantly, its in the area of minaturization where the greatest deviation might be expected to occur. Most of Moore's law in chips has come from continually packing greater numbers of smaller transitors on a chip. Simply printing the designs was one challenge to overcome; with finer designs comes a need for photolithography with wavelengths shorter than visible. This is clearly in the category of problems with Ion Torrent can count as solved by the semiconductor industry.

A second problem is that smaller features are less and less tolerant of smaller and smaller defects in the crystalline wafer of which chips are fabricated from. Indeed, in memory chips the design carries more memory than the final chip so that some can be sacrificed to defects; if excess memory units are left over after manufacturing they are shorted out in a final step. Chips with excessive defects go in the discard bin, or sometimes allegedly are sold simply as lower grade memory units. One wonders if Ion Torrent will give all their partial duds to their methods development group or perhaps give them away in a way calculated to give maximal PR impact (to high schools?).

But, there are other problems which are quite different. For example, with chips a challenge at small feature sizes is that the insulating regions between wires on the chip become so narrow as to not be as reliable. Heat is another issue with small feature sizes and high clock speeds. These would seem to be problems the semiconductor-industry won't pass on to Ion Torrent.

On the other hand, Ion Torrent is trying to do something very different than most chips. They are measuring a chemical event, the release of protons. As the size of the sensor features decrease, presumably there will be greater noise; at an extreme there would be "shot noise" from simply trying to count very small numbers of protons.

Eventually, even the semiconductor industry will hit a limit on packing in features. After all, no feature in a circuit can be smaller than an atom in size (indeed, a question I love to ask but which usually catches folks off-guard is how many atoms are, on average, in a feature on their chip). One possible route out for semiconductors is to go vertical; stacking components upon components in a way that avoids the huge speed and energy hits when information must be transferred from one chip to another. It is very difficult to see how Ion Torrent will be able to "go vertical".

None of this erases that Ion Torrent will be able to leverage a lot of technology from chip manufacturing. But, it will not solve all their challenges. The real proof, of course, will be in Ion Torrent regularly releasing new chips with greater densities. An important first milestone is the on-time release of the second generation chip this spring, which is touted as generating four times as many reads (at double the cost). Rothberg is claiming to have a chip capable of a single human exome by 2012; assuming 40X coverage of a 50M exome, that would require a 200-fold improvement in performance, or nearly four quadruplings of performance. Some of that might come from process or software improvements to increase the yield per chip of a given size (more on the contest in the future); indeed, to meet that schedule in 2 years would insist on either that or a very rapid stream of quadrupling gains.

As I have commented before, they might even pick a strategy where some of the chips trade off higher density for read length or accuracy. For example, applications requiring counting (expression profiling, rRNA profiling) of tags which can be distinguished relatively easily (or the cost of some confusion is small) might prefer very high numbers of short reads.

First of a Torrent?

2011-01-02T09:06:00.002-05:00

For the New Year I've resolved to be a bit more regular in posting here, and lik all New Year's resolutions it is easy to start out big, so there may be a flurry of posting this week. Of course, the real challenge will be to maintain that energy across an entire year. But, to kick-start things I spent the holiday weekend drafting nearly a week's worth of output.

Ion Torrent continues to attract a lot of attention, though its launch last year hasn't yet resulted in my getting hands or eyes on one. Ideally, an evaluation machine would show up but that's happening only in my dreams. Nor did my attempt to win a free one succeed, though one winning entry was of very similar concept (and both were from Massachusetts!). Most of the press has continued to edge towards breathless and unthinking hype, but the counterpoint is in Nick Loman's well-thought bit of exasperation with that hype.

My own thoughts continue to lie in between. I continue to be frustrated by the absurd hype in various tech press outlets, but I also see this as a useful machine. There's a number of interesting angles, which I've decided to tackle with a small series rather than one big lump. Ideally this splitting will result in more coherent arguments on my part, but that's for you to decide.

To me the most frustrating angle is the view that sequencing is a monolith and a single race, with one winner. For Sanger sequencing, this tended to be the case because the underlying technology was so similar and the various platform makers didn't separate much. ABI took the lion's share of the market, Amersham was a distant second and that was almost it. LiCor had the one somewhat differentiated entry, with a different dye system yielding longer reads, but at the cost of greatly reduced throughput. Even these reads were not so much longer (I think they claimed just over a kilobase, whereas ABI routinely got about 3/4 kilobase) to really drive a big niche.

But second-gen has evolved in a very different way. Speed, upfront cost, running cost, library prep, pre-sequencer prep, accuracy and read length are multiple variables in which the different platforms have landed in different boxes. Some of this is inherent in the technologies, whereas others are due to simply design choices or intellectual property positions. An example of the latter is Illumina's patent lock on bridge PCR, whereas other amplification-requiring platforms appear to nearly all use emulsion PCR (Complete Genomics uses rolling circle).
So, to me the question in evaluating a platform and where it is going depends on looking at that particular combination and asking what applications work best. Once that's worked out, the size of the market can be speculated on as well as who else might be bumping elbows in that space.

Now, Ion Torrent has a number of operational features worth noting. First, it has the lowest upfront cost of a sequencer at around $100K fully loaded (sequencer, server & emPCR robots). This is a first point of my annoyance with many glowing articles: they parrot the "$50K" price which buys you just the sequencer. Even worse are the ridiculous claims of Ion Torrent being 1/10th the price of the competition; this is comparing only to the highest price alternative offerings and not the likely alternate choice.

Second, the run times are quite fast. But again, many of those enamored with the device mindlessly spout the time to acquire data and not all the up-front prep. Some of that prep will depend on the particular application, but it is still on the order of 2-3 days to go from DNA sample to data off the sequencer. Now, I know the pain of anticipating data, having recently gnawed my nails off waiting for a high-stakes paired end SOLiD 4 run (closer to two weeks than one), but the truth is a number of other platforms offer similar speed (more in another installment).

Third, the initial release is claiming about 100,000 reads of 100 bp or more (up to about 200). The chip costs $250 and there is another $250 to prep the sample; it is unclear from anything I've seen what is included in that prep cost and in particular how many runs you can get from one such prep. For example, if that includes library adapters and I'm using a direct PCR approach, then that $250 cost is actually inflated. More importantly, if I need more than 100K reads for an application, does that $250 or prep buy me more than one run (i.e. will 200K reads from one sample cost me $750 or $1000?). Error rate is not clear and homopolymers will be a problem, though the probability of miscalling these isn't well documented.

Given these fuzzy estimates, what sort of applications will be best for the Ion Torrent platform in its initial state? To me, and clearly to others, the sweet spot is sequencing of targeted and easily interpetable regions. The two U.S. contest winners (was the European giveaway ever executed?) are just along those lines.

A group at MGH is planning to perform PCR-based targeted sequencing of cancer. This is a very appropriate application which fits many of properties of Ion Torrent. Many cancer mutations are what we call "hotspot mutations"; the same mutations are seen repeatedly. For example, in the very important KRAS oncogene the vast majority of mutations occur in any of the six nucleotides of two adjacent codons. Design your PCR assay correctly, and all you would need is a six base pair readlength (indeed, several tests approved for the clinic or on their way there could be seen as 1-bp read length sequencing assays). More realistically, you need to set the primers back a bit from the hotspot and read through the primers, but for this the 100 bp reads of Ion Torrent will be quite good. Now, this hotspot behavior governs most, but not all, activating mutations in oncogenes. This can be seen as it being hard to turn something on by tinkering with it, though in a few cases the tinkering is by removing a whole inhibitory exon and there are many ways to do that. On the other hand, many tumor suppressors are mutated in a diffuse pattern. Sometimes there are hotspots due to particular mutation processes or other forces, but these are never as hot.

The other winning entry was from Woods Hole Marine Biological laboratory to rapidly profile to identify bacterial contamination of water. Again, PCR-based and looking at well-defined signatures, in this case ribosomal RNA profiles.

Each example fits well into the Ion Torrent's capabilities. In both cases, you don't need enormous numbers of reads to do a decent job, though more reads would let you either look for rarer species or assay more loci. Since you are looking for signatures, the assays can be calibrated well in advance versus the error and read length characteristics of the platform. For example, you can know in advance where there are homopolymer runs and adapt for them. Offhand, I can't think of an oncogenic hotspot that involves a homopolymer run and in oncogenes the frame must generally be preserved (again, there are those rare non-coding oncogenic changes) so that would help constrain errors.

Given that sweet spot who is going to feel Ion Torrent's elbows? The obvious candidate is Roche. The 454 GS Jr is around 2-3X the upfront cost (again, for a complete infrastructure), around 4X the cost per run and will yield 0.5-1X the number of reads -- but much longer ones. However, for both the applications above long reads aren't really such a great advantage. Again, for many of your signatures you can design the signature around the read length, and really long reads add only a bit more value. For dealing with clinical cancer samples, you really want to keep your PCR amplicons down to 250bp or less because the DNA you get is generally quite fragmented and has other impediments to PCR. With a protocol that can read in from each end of the PCR fragments (perhaps randomly or perhaps in two separate runs, one from each end), Ion Torrent's current length fits well. Short signatures will work better on both platforms in any case, as in the real world you get some reads that peter out much sooner -- short signatures mean more effective reads of a signature per run. 454 has a more established chemistry and performance specs, but I would expect Ion Torrent to be serious competition for the Jr platform, with the 454 family holding on to the applications (such as HLA haplotyping) where length really does matter. Holding on, that is, until Ion Torrent can push their read lengths to similar territory.

That's a pretty big lump. Next installment (not necessarily tomorrow; I might interleave some other topics burning on my desk), looking at the much stressed tie to the semiconductor industry.

Google's Ngram Viewer

2010-12-20T23:19:00.002-05:00

I've been playing off and on with Google's Ngram viewer since it was announced on Friday. This is the tool that enables you to graph the frequency over time in usage of given words or phrases. All sorts of interesting experiments are possible -- for example, try comparing the usage of a word and a synonym vs. an antonym or a euphemism to compare their usage (or, you could examine those three words -- "antonym" seems to be much less frequently used but growing in frequency!).

But, I've already noted some anomalies. The plot for "United States of America" is surprisingly spiky, with surprisingly few mentions in the early 1800s. That is perhaps an artifact of the sources available for the Google book digitization project, but it does cast concern on some of the conclusions being drawn from this tool.

But worse, there are definitely some issues with dating and with automated text recognition. Search for "Genomics", and some awfully early references show up. These seem to fall into two categories: serious book dating errors and text errors. In the former category, I don't believe Nucleic Acids Research published in 1835, and a number of other periodicals seem to be afflicted with similar misdatings. In the latter, "générales" seems to be a favorite to transmute to "genomics".

These issues do not invalidate the tool, but they do urge caution in interpreting results -- particularly if trying to explore the emergence and acceptance of a new term.

An approach to deal with this would be to turn the problem around. A systematic search for anachronistic word patterns could identify misdatings or questionable datings in either direction. Not only would this identify documents transported backwards in time, but also ones which should be flagged for time travel in the other direction. For example, using the tool I discovered that someone sharing my surname co-authored a screed against Masonry back in the 1700s -- and this same work shows up as a modern book due to a reprinting in recent years.

But in any case, it is an interesting way to explore language and culture. Even without a little tidying & curation.

Bone Marrow Registries of Contention, and the Future of Tissue Typing?

2010-12-19T21:40:00.003-05:00

This summer I took TNG to the Portsmouth Air Show to enjoy viewing aerobatics and looking at some aircraft up close. As with many such events, there is a vendors section & at this one I came across the Caitlyn Raymond Bone Marrow Registry. Curious to check out someone else's consent form, experience a buccal swab & contribute to a good cause, I signed up. Quick & painless. It's also only the second time I've consented to have my own DNA analyzed, and the first professional job (we sequenced one polymorphism from each student in an undergrad class at Delaware).

I don't regret that decision, but one part that was a bit odd at first was filling out my medical insurance information. Okay, someone has to pay for the DNA testing but it seemed a little odd to stick my insurance with it -- but I didn't give it a lot of thought at the time. Since that time, I've regularly seen the registry at various community events as well as a kiosk at the local mall.

Yesterday's Globe had an article causing me to revisit that memory. U Mass Medical Center runs the Caitlyn Raymond registry, and someone there saw a dubious opportunity and ran with it. The lead for the article focused on the fact that professional models had been used as greeters at many events, helping a very high recruitment rate. Okay, that could be seen as just creative. But, the back end is that U Mass has been charging as much as $4K per sample for testing. YIKES! That's in excess of what I've heard BRCA testing goes for. Now U Mass will be getting a lot of attention from a number of attorneys general.

One point in the article is a concern that the use of models may compromise the informed consent process. The proof, as it continued, will be if registrants from the Raymond pool fail to follow-through with donations at an unusually high rate, but given that most will never be contacted it may never be known.

But it got me thinking: since the testing is purely a DNA analysis, then presumably each complete human genome sequence can be used to type an individual. Perhaps even the tests from 23 et al* hit the right markers, or at least some tightly linked ones.

So, is it ethical to reach out to such individuals? Given that I could, in theory, search the released DNA sequences from the personal genome project, would it be reasonable to try to track one down and beg for a donation? Of course, the odds of a successful match are tiny -- but as more and more PGP sequences pile up, the chance of such a search succeeding go up.

What about non-public DNA databases? Again, suppose a 23 et al had the right markers (or nearly so). Should you have to opt-in to be notified that you are predicted to be a possible donor match? Is there a mechanism to publish profiles to a central database, with an ability to ping the user back if a match is made? And if every newborn is being typed for a few thousand other markers, will testing for transplantation markers also be required?

* -- a great term, I believe originated by Kevin Davies in his $1K genome book.

Is Pacific Biosciences Really Polaroid Genomics?

2010-12-10T21:07:00.004-05:00

The New England Journal of Medicine this week carries a paper from Harvard and Pacific Biosciences detailing the sequence of the Vibrio cholerae strain responsible for the outbreak of cholera in Haiti. The paper and supplementary materials (which contains detailed methods and some ginormous tables) are free for now. There's also a nice piece in BioIT World giving a lot of backstory. Not a few other media outlets have carried it as well, but that's where I've read.

All in all, the project took about a month from an initial phone call from Harvard to Pacific Biosciences until the publication in NEJM. Yow!! Actual sequence generation took place 2 days after PacBio received the DNA. And this is sequencing two isolates (which turned out to be essentially identical) of the Haitian bug plus three reference strains. While full sequence generation took longer, useful data emerged three hours after getting on the sequencer (though there are apparently around 10 wall clock hours of prep before you can get on the sequencer). With the right software & sufficient computational horsepower, one really could imagine standing around the sequencer and watching a genome develop before your eyes (just don't shake the machine!).

Between this & the data on PacBio's DevNet site (you'll need to register to get a password), in theory one could find the answers to all the nagging questions about the performance specs. Actually, this dataset is apparently available as the assembled sequence but only summary statistics for certain aspects of it. For example, apparently dropped bases are focused on C's & G's, so these were discounted.

Read lengths were 1,100+/-170bp, which is quite good -- and this is after filtering out lower quality data -- and 5% of the reads were monsters bigger than 2800 bases. It is interesting that they did not use the circular consensus method, which was previously published in a method paper (which I covered earlier) and yields higher qualities but shorter fragments. It would be particularly useful to know if the circular consensus approach effectively dealt with the C/G dropout issue.

One small focus of the paper, especially in the supplement, is depth of sequence analysis to infer copy number variation. There is a nice plot in Supplementary Figure 2 illustrating how the copy number varies with distance from the origin of replication. If you haven't looked at bacterial replication before, most bacteria have a single circular chromosome and initiate synthesis starting at one point (the 0 minute point in E.coli). In very rapidly dividing bacteria, the cell may not even wait for one round of synthesis to complete before firing off another synthesis round, but in any case in any dividing population there will be more DNA near the origin than near the terminus of replication. Presumably one could estimate the growth kinetics based on the slope of the copy number from ori to ter!

After subtracting out this effect, most of the copy number fits a Poisson model quite nicely (Supplementary Figure 3). However, there is still some variation. Much of this is around ribosomal RNA operons, which are challenging to assemble correctly since they appear in arrays of nearly (or completely) perfect repeats which are quite long. There's actually even a table of the sequencing depth for each strain at 500 nucleotide intervals! Furthermore, Supplementary Figure 4 shows the depth of coverage (uncorrected for the replication polarity effect) at 6X, 12X, 30X and 60X coverage, illustrating how many of the trends are actually noticeable in the 6X data.

What biology came out of this? A number of genetic elements were identified in the Haitian strains which are consistent with it being a very bad actor and also that it is a variant of a nasty Asian strain.

All-in-all, this neatly demonstrates how PacBio could be the backbone of a very rapid biosurveillance network. It is surprising that in this day-and-age that the CDC (as detailed in the BioIT article) even bothered with a pulsed field study; even on other platforms the turnaround for a complete sequence wouldn't be much longer than to do the gel study, and the results are so much richer. Other technologies might work too, but the very long read lengths and fast turnaround offered should be very appealing, even if the cost of the instrument (much closer to $1M than to my budget!) isn't. But, a few instruments around the world serving other customers but with priority given to such samples could form an important tripwire for new infections, whether they be acts of nature or evil persons. Now, it is important to note that this involved a known, culturable bug and the DNA was derived from pure cultures, not straight environmental isolates.

On a personal note, I am quite itchy to try out one of these beasts. As a result, I'm making sure we stash some DNA generated by several projects so that we could use them as test samples. We know something about the sequence of these samples and how they performed with their intended platforms, so they would be ideal test items. None of my applications are nearly as exciting as this work, but they are the workaday sorts of things which could be the building blocks of a major flow of business. Anyone with a PacBio interested in collaborating is welcome to leave me a private comment (I won't moderate it through), and of course my employer would pay reasonable costs for such an exercise. Or, I certainly wouldn't stamp "return to sender" on the crate if an instrument showed up on the loading dock! I don't see PacBio clearing the stage of all competitors, but I do see it both opening new markets and throwing some serious elbows against certain competing technologies.

Arsenic and New Microbes

2010-12-03T23:06:00.002-05:00

Yesterday's announcement of a microbe which not only tolerates arsenic but actually appears to incorporate it in place of phosphorous has traveled a typical path for such a discovery: while it is quite a find, the media has generated more than a few ridiculous headlines. Yes, this potentially expands the definition of life, at least in an elemental sense, but it hardly suggests that such life forms exist elsewhere. A similar absurd atmosphere briefly reigned around a discovery of a potentially habitable world around a distant star -- the discoverer was quoted in at least one outlet that his find was guaranteed to have life. Given that we know very little about the probability of life starting, I always cringe when I hear someone announce that such events are either certain or certainly impossible; we simply can't calculate believable odds given our poor knowledge base. On the other end, suggestions have been raised as to this bug being a starting point for bioremediation of arsenic-contaminated aquifers; but really this discovery isn't a huge step in that direction beyond species already known to tolerate the stuff. It's also disappointing that none of the popular news items I've seen have pointed out how a periodic table can be read to show chemical similarity of phosphorous and arsenic.

That said, it is an intriguing discovery. The idea that all those phosphates on the metabolic diagrams might be substituted with arsenate is quite jarring. No reader of this space will be surprised to hear me advocate for immediate sequencing of this bug (if it hasn't already happened and just not yet reported). A microbial genome these days can be roughed out in well under a month (actually, sequence generation for Mycoplasma a decade ago took that long; clearly we can go faster now).

In order to interpret that genome, though, another whole line of experiments is needed. Assuming that the ability of this organism to incorporate arsenate in place of phosphate is confirmed, some of the precise enzymes capable of doing this trick need to be located. Simply finding arsenate-analogs of some key metabolites (such as phosphorylated intermediates in glycolysis) would point at a few enzymes, and then it would be valuable to demonstrate the purified enzymes pulling the trick. The next step then would be to test whether more conventional enzymes have this activity. Despite what many of us learned in various exposures to biochemistry from elementary school on up, enzymes aren't utterly specific for their substrates. Instead, there is a certain degree of promiscuity, though generally not with equal activity. So, to extend my analogy, if the new bug's triose phosphate isomerase can work on triose arsenates, then testing that activity in well-characterized TPIs would in order.

Assuming that such enzymes (from E.coli or human or yeast or what-not) do not have the activity, then crystal structures of the arsenate-lover would be an important next step. Of course, repeating this for the whole roster of enzymes in the bug would be quite an undertaking, but perhaps a number could be modeled to see if a consistent pattern of substitutions or other alterations emerges.

At one time, it was vogue to speculate on life forms which used silicon in place of carbon, given it's location one rung down on the periodic table. Did any author ever dare suggest arsenic for phosphate? I doubt it, but perhaps there was some mind playing with the possibilities who wrote it down somewhere (along with a large pile of other guesses that will not pan out).

Mild alleles in severe diseases: an opportunity for enlightenment

2010-11-03T08:44:00.002-04:00

Monday's Globe had a blurb about a book signing which re-kindled a previus interest of mine. The author, Michael Dana Kennedy, had quit his job as a medical researcher to write the novel, a tale of two brothers ending up on opposing forces in the Pacific during WW2. While the book concept might have enough appeal to go to the back of my infinite reading list, it's the author's backstory that really grabs me.

The reason Kennedy quit his job is that it was perceived as a health threat: he was diagnosed with cystic fibrosis in his mid-50s. This reminded me of an elderly female patient the Gene Sherpa had mentioned who he had diagnosed with CF.

Cystic fibrosis is a very difficult disease, and for a long time few patients made it out of their twenties. I understand that with modern care, including antibiotics and regular respiratory therapy, many patients live substantially longer. But, this underscores what I find interesting about these two patients -- without any treatment at all they have long outlived most of their peers afflicted with cystic fibrosis. Hence, they must have comparatively mild cases. And those should be interesting.

The key question is why are these cases so mild? The simplest answer would be that they carry at least one allele which retains substantial function of CFTR (the gene mutated in CF). The more complex answer would be that they carry other genetic variants which substantially moderate the impact of the defective allele(s). Either answer would be very enlightening, both for CFTR specifically and for better understanding protein function in general.

I would expect that with the PGP and 1000 genomes project and all the other human genome sequencing efforts public and private, many new alleles will be discovered in many well understood disease genes (or as well understood as any disease gene is). A key follow-up to execute, when possible, is to determine which of these alleles had health impact. CF is appealing from this angle because we know a biochemical phenotype (altered salt excretion) which can be measured and we know a possible medical issue to assess (history of frequent respiratory infections). BRCA1 would be another valuable case where we already know many disease alleles, though there the question is more complicated to answer. I'm sure there are many more.

Studying some of these easier cases will, with luck, help shed some light on the avalanche of novel genetic variants which are pouring from germline genome projects -- and an order of magnitude higher from cancer genome projects (since in many tumors there is some combination of deficient DNA repair/replication as well as significant historical exposure to mutagens such as cigarette smoke). Lacking good high-throughput ways to assess most of these functionally, it would behoove the community to leverage the ultimate functional tests -- human survival.

Plenty of Genomes are Still Fair Game for Sequencing

2010-10-31T21:36:00.004-04:00

I've been grossly neglecting this space for an entire month with only the usual excuses -- big work projects, a lot of reading, etc. None good enough. Worst of all, as usual, it's not that I haven't composed possible entries in my head -- they just never get past my fingertips.

Tonight is the night most associated with pumpkins, and an earlier highlight was attending the Topsfield Fair, where the pictured specimen was on display. Amazing as it is, it fell nearly 15 pounds shy of the world record. If you want to try to grow your own, every year the variety which has dominated the winners can be purchased. Nature isn't all though; champion pumpkin growing requires a lot of specialized culture ranging from allowing only a single fruit to set to injecting nutrients just upstream of that fruit.

Sometime in recent memory there were some other blogs noted in GenomeWeb for discussing whether there are any truly remarkable genome sequencing projects left. Which I've been pondering: what makes for a very interesting species to sequence. Now, both of the bloggers mentioned clearly were not fond of either "K" genome project -- the 1,000 humans or 10,000 vertebrates. There were also some potshots taken at the "delicious or cute" genomes concept. One suggested that no interesting metazoa ("animals") are left.

So, what does make an interesting genome? Well, I can think of several broad categories. I'll try to throw out possible examples of each, though to be honest I wouldn't be surprised if some of these genomes are sequenced or nearly so -- it's very hard to keep track of complete genomes these days!

First, which I think would resonate with those two critical articles, would be genomes with interesting histories -- genomes that might tell us stories purely about DNA. This was the bent of these papers I refer to. In particular, they were thinking of many of the unicellular eukaryotes which are the result of multiple endosymbiont acquisition / genome fusion events. But, I would definitely throw into this category a particular animal: the Bdelloid rotifers, which have gone without recombination for a seeming eternity. Of course, to really understand that genome, you'd need to also sequence one of the less chaste rotifers.

Another hugely interesting class of genomes would be those to shed light on development and its evolution (evo-devo). In particular, there are a lot of arthopod genomes yet unsequenced -- from what I've noted it appears that most sequenced arthropods are either disease vectors, agricultural pests or economically important (plus, of course, the model Drosophila). Even so, I'd guess there are not many more than a dozen complete arthopod genomes so far -- quite a paucity considering the wealth of insects alone. And, if I'm not mistaken, mostly insects and an arachnid or two have gone fully through the sequencer -- where are all the others? By the way, I'd be happy to help with sample prep for the Homarus americanus genome!

Another huge space of genomes worth exploring are those were we are likely to find unusual biochemistry going on. Now, a lot of those genomes are bacterial or fungal, but there are also an awful lot of advanced plants that have interesting & useful biochemical syntheses.

All that said, I find it odd that some don't see the import and utility of sequencing many, many humans and a lot of vertebrates also. It is important to remember that a lot of funding is from the public, and the public considers many of these other pursuits less important than making medical advances. It is easy for those of us in the biology community to see the longer threads connecting these projects to human health or just the importance of pursuing curiosity, but that doesn't always sell well in public.

An optimistic view is that all the frustrated sequencers should hunker down and patiently wait; data generation for new genomes is getting cheaper by the minute, with short reads to fill out the sequence and ultra-long reads to replace physical mapping. A more conservative view holds that bioinformatics & data storage will soon dominate the equation, which might still make it hard to get lots of worthy genomes sequenced.

Personally, I can't stroll a country fair without wanting to sequence just about everything I see on display -- the chickens that look like Philadelphia Mummers, the two yard long squash, bizarrely shaped tomatoes -- and of course, the three quarter ton plus pumpkins.

Scenes from the Cancer Personalized Medicine Wilderness

2010-09-28T20:40:00.004-04:00

I'm going to attempt to synthesize a number of thoughts which I've long pondered along with a bunch of news items I came across today. With luck, the result will be coherent and I'll not make a fool of myself.

There was a very interesting article last week in the New York Times on a serious ethical dilemma in melanoma and how different specialists in the field are voicing opinions on both sides of the divide. Even better, today I came across an excellent blog post reviewing that article which also added a lot of expert background. I'll summarize the two very quickly.

Metastatic melanoma is an awful diagnosis; the disease is very aggressive. Furthermore, the standard-of-care chemotherapy drug is a very ugly cytotoxic, with nasty side effects and very poor efficacy (more on that later). Sequencing studies have revealed that well over half of metastatic melanomas have a mutant form of the kinase B-RAF (gene: BRAF), most commonly the mutation V600E (which, alas, due to some sequencing error was for a while known as V599E). That's the substitution of an acidic residue (glutamate) for a hydrophobic one (valine), and it is right in the kinase active site.

Now, a biotech called Plexxikon, in conjunction with Roche, has developed an inhibitor of B-RAF called PLX4032. In Phase I trial results reported this summer in the New England Journal of Medicine, very promising tumor regressions were seen. Now remember, this was a single-arm Phase I trial for safety, meaning we don't have an objective comparison to make.

And there begins the rub. To some doctors (and many patients), the combination of great preclinical results, the theoretical and experimental underpinnings for targeting B-RAF in melanoma and the observed regression means we have a winner on our hands and it is now unethical to have a randomized trial comparing the new compound against the standard-of-care.

At the other pole are doctors who worry that we have been fooled before.
My standard example to trot out for such cases is a famous CAST cardiovascular trial to which a placebo arm was grudgingly added -- a sound theory had been advanced that
suppressing arrythmias in certain patients would prevent death. CAST was stopped early when it was clear the placebo arm fared far better; the toxicities of the drugs overwhelmed any benefits. Even closer to our current story is the drug sorafenib, which was originally developed as a B-RAF antagonist. Now, there are many in the field who argued that it really wasn't, but Bayer and Onyx got it to market (probably based on its inhibition of numerous other kinases) and the "raf" syllable in the generic name points to their belief in the B-RAF theory. Unfortunately, in randomized clinical trials it failed to work in V600E melanomas.

One idea that was apparently floated by at least one oncologist working in the trials, but rejected by the corporate sponsors, was to try to win approval based on nearly miraculous recoveries seen in some patients on death's door. What the NYT article failed to discuss is whether the FDA would buy that argument; there are many reasons to think they wouldn't -- they really do not like single arm trials, because all too often spurious results occur do to random chance (or rarely, to manipulation of the trial).

An important idea discussed in all this is the concept that once we have established a therapy as efficacious, it is generally unethical to withhold that therapy from patients. But, we are often not on such solid ground even in this area. Clinical trials represent a horrible case of multiple testing; more than a few drugs that squeaked through their trial would not if you ran the trial again; they just got lucky. Don't believe me? Think back to Iressa, which received accelerated approval for lung cancer and then had it withdrawn (only to later be reintroduced). We now know a key piece of that particular puzzle: Iressa works in patients whose tumors have mutant forms of the EGFR. The first trial, by chance, was enriched for such patients and the second trial (also by chance) was not as enriched. Given that the EGFR hypothesis wasn't known, neither trial could have been manipulated.

But another recent item, covered in a different post on the same blog, reminds us that even well-established clinical approaches may not hold true over time. Screening mammography is a hot potato issue in cancer: can you save lives by screening healthy women for breast cancer. Various studies have tried to ask this question not just for women overall, but by age groups since the incidence of breast cancer and the quality of mammograms changes with patient age. The newest fuel on this fire is a very clever Norwegian study, which I won't attempt to summarize, that suggests that much (but perhaps not all) of the benefit of screening mammography has been eroded by improvements in cancer care. In other words, the advantage of early detection has been blunted by better treatments. Now, I'm not qualified to really review that study, but certainly this is a concept we should keep in mind: the utility of medical strategies may change over time, and not always for the better.

In my mail tonight was a thick magazine-sized volume from Scientific American, which I confess I am not a subscriber of (it's a fine magazine; I just already subscribe to too many fine magazines). This special edition, titled "Pathways: The changing science, business & experience of health", focuses on healthcare with a mix of articles. Some appear to be written by professional writers, while others are thinly-veiled advertisements for various companies.

In scanning the table of contents, I was caught by "Pioneering Personalized Cancer Care", though unfortunately this turns out to be one of the puffier pieces. Written by two principles in the company, it mostly describes N-of-one, a company which has as its customers cancer patients. N-of-one tries to distill the available knowledge on a person's tumor and help them navigate to the most appropriate tests. It's a business model I've sometimes wondered about for myself, since playing an oncologic Sherlock Holmes could be both fascinating and rewarding. On the other hand, the regulatory environment is fraught with uncertainty and most likely this sort of organization will have to rely on wealthy customers willing to pay their own way.

Now, the article did set my teeth on edge early on with the statement "Recently, projects such as the Cancer Genome Atlas have documented thousands of mutations in cancer cells that can lead to unregulated cell growth and prevent apoptosis (cell death), the hallmarks of malignancy". Any regular reader of this space knows that I am a gung-ho proponent of sequencing tumors, but with that comes an obligation to be honest. And the honest truth is that sequencing has yielded thousands of candidates, but only a handful of those have actually been shown to have transforming ability -- there's just no high-throughput way to do that en masse.

But, what N-of-one and others are doing is where I strongly believe the future of oncology lies. But, it will be a complicated place. Getting back to B-RAF, I've heard noise that it has been found in a number of additional tumor types, albeit at low frequency. So, supposes it occurs at 1 in 1000 frequency in some awful tumor type. With routine whole-genome sequencing of tumors, we could detect that. Such sequencing is starting to be used to good effect, as reported recently in Nature. That leads to a conundrum for everyone. For a patient or clinician, do you go with PLX4032, given that we know it targets BRAF -- but knowing that we don't know whether BRAF is really driving your tumor (especially if the mutation is not V600E)? For those wanting to design clinical trials, could you really find enough patients to stock a trial -- or are you willing to have a trial with "any cancer, as long as it has a BRAF mutation"?

This is the challenge that personalized medicine presents us. With genome sequencing (and eventually also routine whole methylome profiling), we can find what makes cancers different -- but how will we ever actually sort through all those differences? Should we move away from randomized trials to going where the science seems to lead us, even knowing that more than a few times there have been dead ends?

I can find only one easy answer to all this: don't trust anyone who offers an easy answer to all this.

Review: The $1000 Genome

2010-09-21T23:18:00.005-04:00

Kevin Davies' "The $1000 Genome" deserves to be widely read. Readers of this space will not be surprised that there are a few changes I might have imposed had I been its editor, but on the whole it presents a careful and I think entertaining view of the past and possible future of personal genomics.

The book is intended for a far wider audience than geeky genomics bloggers, so the emphasis is not on the science. Rather, it is on some of the key movers-and-shakers in the field and some of the companies which have been dominating this space, ranging from the first personal genetic mapping companies (23 and Me, Navigenics, Pathway Genomics and deCodeMe) to the instrument makers (such as Solexa/Illumina, Helicos, Pacific Biosciences, ABI and Oxford Nanopore) to those working on various aspects of human genome sequencing services (such as Knome and Complete Genomics. Various ups and downs of these companies -- and the debates they have engendered -- are covered as well as the possible impacts on society. Along the way, we see a few glimpses of Davies exploring his own genome and some of the biological history which he seeks to enlighten through these expeditions.

It is not a trivial task to try to explain this field to an educated lay public, but I think in general Davies does a good job. The overviews of the technologies are limited but give the gist of things. Anyone writing in this space is faced with the dilemma of trying to explain too much and losing the main thread or failing to explain and preventing the reader from finding it. Mostly I think he has succeeded in threading this needle, perhaps because only rarely did I feel he had missed. One example I did note was in explaining PacBio's technology; hardly anyone in science will know what a zeptoliter is, let alone someone outside of it. On the other hand, what analogy or refactoring of that term could remove it from the edges of science fiction? Not an easy challenge!

For better or worse, once I've decided I generally like a book like this my next thoughts are what could be removed and what could be added. I really could find little to remove. But, there are a few things I wish were either expanded or had made it in altogether.

It would be dreary to enumerate every company which has ever thrown its hat in the DNA sequencing ring. It is valuable that Davies covers a few of the abject failures, such as Manteia (which did yield some key technology to Illumina when sold for assets) and US Genomics. There is scant coverage, other than by mention, of most of the companies which have but nascent attempts to enter the arena. However, the one story I really did miss was anything about the Polonator. It's not that I really think this system will conquer the others (though perhaps I hope it will hold its own), it just represents a very different tack in corporate strategy that would have been interesting to contrast with the other players.

Davies has been in the thick of the field as editor of Bio IT World, so this is no stitching together of secondary sources. I also appreciated that he includes both the ups and the downs for these companies, emphasizing that this has not been easy for any of them. But, that added to my surprise at several incidents which were left out (believe me, many were left in I had never heard before). Davies describes how Helicos delivered an instrument to the CRO Expression Analysis, but not that it was very publicly returned for failing to perform to spec. Nor is Helicos' failed attempt to sell themselves mentioned. An interesting anecdote on Complete Genomics is how a wildfire nearly disrupted one of their first human genome runs; left out is the near-death experience of that company when it was forced to either lay off or defer salaries for nearly all of its staff. The section on Complete's founder Rade Drmanac mentioned Hyseq, but not the company (or was it two) which he ran between Hyseq and Complete to try to commercialize sequencing-by-hybridization. This would have added to this portrait of determination -- and the travails of the corporate arena. I was also surprised that the short profile of Sydney Brenner as a personal genomics skeptic didn't include the fact he invented the technology behind Lynx, which was another early attempt in non-electrophoretic sequencing. Some would see that as irony.

Another area I would like to have seen expanded was the exploration of groups such as Patients Like Me, which are windows on how much people are willing to chance disclosing sensitive medical information. One section explores the fact that several prominent persons interested in this field became so when their children were diagnosed with rare recessive disorders, leading them to ponder whether they would have made the same marriage had they known in advance of this danger. I was surprised that little of the existing experience in this area was explored; I believe the Ashkenazi population has dealt with this in screening for Tay-Sachs and other horrific disorders which are prevalent there.

The book is stunningly up-to-date for something published the beginning of September; some incidents as late as June are reported. Despite this, I found little evidence of haste. I'm still trying to figure out what a "nature capitalist" is, but that's the only case I spotted of a likely mis-wording.

Davies briefly explores possible uses of these sequencing technologies beyond our germline sequences, but only very briefly. Personally, I think that cancer genomics will have a more immediate and perhaps greater overall impact on human medicine, and wish it had gotten a bit more in depth treatment.

Davies in a expatriot Brit, living not very far from me. The sections on the possible impact of widespread genome sequencing on medicine are written almost entirely from a U.S. perspective, with our hybrid public-private healthcare system. I suspect European readers would hunger for more discussion of how personal genomics might be handled within their socialized medical systems and different histories of handling the ethical issues (Germany, I believe, has pretty much banned personal genomics services). On this side of the pond, he does a nice job of showing how different state agencies have charged into the breach left, until recently, by the FDA.

Okay, too many quibbles. Well, maybe one last one -- it would have been nice to see more on some of the academic bioinformaticians who have created such wonderful and amazing open-source tools as Bowtie and BWA.

As I mentioned above, Davies injects a good amount of himself into all this. I've encountered books (indeed, on recently on moon walkers), in which this becomes a tedious over-exposure to the author's ego. This is not such a book. The personal bits either link pieces of the story or make them more approachable. We find out that he has already attained a greater age than his father did (due to testicular cancer, one of the few cancers in which overwhelming progress has been made), leading to questions he hopes his genome can answer. Hence, his trying out of pretty much all of the array-based personal genetic services. But, he does not address one question that the book raised in my mind: will the royalties from this project fund a complete Davies genome?

ARID1A A Fertile Ground for Mutations in Ovarian Clear Cell Carcinoma

2010-09-11T08:12:00.009-04:00

Although ovarian clear cell carcinoma does not respond
well to conventional platinum–taxane chemotherapy
for ovarian carcinoma, this remains
the adjuvant treatment of choice, because effective
alternatives have not been identified.

This sentence is a depressing reminder of the status of medical treatment of far too many tumor types. Present in roughly 12% of U.S. ovarian cancer cases, ovarian clear cell carcinoma (OCCC) is a dreadful diagnosis.

Two papers this week made a significant step forward in understanding the molecular basis -- and heterogeneity -- of this horror. Seemingly the finale of an old-fashioned race to publish, groups centered at the British Columbia Cancer Center (in New England Journal of Medicine) and Johns Hopkins University (in Science) published papers with the same headline finding: inactivating mutations in the chromatin regulating gene ARID1A (whose gene product is known as BAF250) are a key step in many -- but not all -- OCCC. I'll use the shorthand Vancouver and Baltimore to refer to the respective groups.

Both papers got here by the largest applications of second generation sequencing to cancer so far published. The Vancouver work relied on transcriptome sequencing (RNA-Seq) of a discovery cohort of 18 patients; the Baltimore group used hybridization targeted exome sequencing on just 8 patients. Both used Illumina paired-end sequencing for the discovery phase; Vancouver also used the same platform for validation on a larger cohort.

Whole genome sequencing is likely the future for cancer genomics. A non-cancer paper just published 20 genomes in one shot, underscoring how this is becoming routine with easy samples & a work which is apparently in press (I have no inside knowledge; it has been discussed at several public meetings) will have perhaps a dozen human genomes in it. But, there are still cost advantages to focusing on expressed genetic regions (and perhaps a bit more) and perhaps further information to be gleaned from actually looking a gene expression. These two papers give an opportunity, albeit a bit constrained, to compare the two approaches.

One interesting note comes straight out of the Vancouver data. After finding ARID1A mutations in 6/18 discovery samples, they re-screened those samples plus 211 additional samples. In total this set included 1 OCCC cell line, 119 OCCC, 33 endometrioid carcinomas and 76 high-grade serous carcinomas. The validation screen was by long-range PCR (mean product size 2067 bp) products sheared and sequenced on the Illumina. One exon proved troublesome and required further PCR and sequencing by Sanger. In any case, the key bit here is in the discovery cohort this approach found ARID1A mutations which had been missed by the original RNA-Seq. As the authors state, a likely culprit is nonsense mediated decay (NMD). It would be interesting to go into their dataset to see if these samples had a markedly lower expression of ARID1A, though I don't have easy access to it (it has been deposited, but with protections that should be the subject of a future post).

One interesting contrast between the two studies is the haul of genes. The Vancouver group found ARID1A as a recurrently mutated gene; the Hopkins group not only bagged ARID1A but also KRAS, PIK3CA and PPP2R1A. KRAS and PIK3CA are well-known oncogenes in multiple tumor types and had previously been implicated in OCCC, but PP2R1A is a novel find. The Vancouver group did specifically search for KRAS and PIK3CA mutants in their cohorts by PCR assays and found one patient sample and one cell line with KRAS mutations. Again, it would be interesting to review the RNA-Seq data to generate hypotheses as to why these were not found in the Vancouver set. On the other hand, the RNA-Seq data did identify one case of a rearranged ARID1A. While it is possible to use hybridization capture to identify gene fusions, this cannot be practically done in a hypothesis-free manner. In other words, without advance interest in ARID1A that approach would not work. In addition, CTTNB1 (beta catenin) mutations had been found previously in OCCC and were specifically checked (and found) by the Vancouver group, but none were reported by the Baltimore group. One final small discrepancy: both groups looked at cell line TOV21G for their mutations of interest and both found the same activating KRAS and PIK3CA alleles. However, Vancouver found one ARID1A allele but Baltimore found that one and a second one (actually, the two mutations I am calling the same [1645insC and 1650dupC] aren't described precisely the same, though I'm guessing it is a difference in an ambiguous alignment).

One other surprise is that TP53 (p53) and PTEN mutants had apparently been reported either for OCCC or endometriosis-associated tumors, yet neither group reported any.

An analysis that is not explicitly found in either paper but I feel is valuable is to look at the co-occurrence of these mutations. If we look only at patient samples, then the big take-home is that neither group saw co-occurrence of KRAS and ARID1A (the TOV21G cell line is at odds with this conclusion). Mutually-exclusive mutations have been seen in many tumors. For example, KRAS mutations are generally mutually-exclusive with other mutations in the RTK-RAS-RAF-MAPK pathway. In contrast, ARID1A mutations are found in conjunction with mutations in CTTNB1, PIK3CA and PPP2R1A -- one patient sample in the Baltimore data was even triple mutant for ARID1A, PIK3CA and PPP2R1A. About 30-40% of sample are mutated for none of these genes as far as this data can tell; the hunt for further causes will continue. Will they be epigenetic? Mutations in regulatory elements?

Another interesting comparison is simply the number of mutations per sample. The Hopkins exome data typically has very small numbers of mutations (after filtering out germ line variants); as few as 13 in a sample and as many as 125 -- and the high number was from a tumor which had previously been treated with DNA-damaging agents (all of the other tumors in the Hopkins study were treatment naive). In contrast, the Vancouver data often found more than 1000 non-synonymous variants per tumor. Unfortunately, no clinical history information is available for the Vancouver cohort, so we don't know if this is from DNA-damaging therapeutics or differences in the sequencing or variant filtering. In an ideal world, we could filter each data set with the other group's filtering scheme to see how much of an effect that would have.

The Vancouver group went beyond sequencing to examine samples by immunohistochemistry (IHC) for expression of the ARID1A gene product, BAF250. There is a strong, but imperfect, negative correlation between mutations and BAF250 expression. Some mutated but BAF250-expressing samples may be explained by the target of the antibody; the truncated forms may still express the correct epitope. Alternatively, ovarian cells may be very sensitive to the dosage of this gene product (in some samples both wt and mutant alleles were clearly found in the RNA-Seq data). Also of interest will be samples lacking expression but unmutated; these may be the places to identify further mechanisms for tumors to eliminate BAF250 expression.

The Vancouver study illustrates one additional bonus from RNA-Seq data: a list (in the supplemental data) of genes differentially expressed between ARID1A mutant and ARID1A wild-type cells.

Another interesting bit from the Vancouver paper is looking at two cases in which the tumor was adjacent to endometrial tissue. In one of these, the same truncating mutation was found in the adjacent lesion and tumor -- but not in a distant endrometriosis. Hence, the mutation was not driving the endometriosis but occurred afterwards.

I'm sure I'm short-shrifting further details from the paper; there's a lot of data packed in these two reports. But, what will it all mean for ovarian cancer patients? Alas, none of the genes save PIK3CA are obvious druggable targets. PIK3CA encodes the alpha isoform of PI3 kinase, a target many companies are working on. But that wasn't novel to these papers. PP2R1A is a regulatory subunit of a protein phosphatase and the mutations are concentrated on a single amino acid, suggesting these are activating mutations (as seen in ARID1A, inactivating mutations can sprawl all over a gene). Phosphatases have not been a productive source of drugs in the past, but perhaps that can be changed in the future. Chromatin regulation is a hot topic, but ARID1A is deficient here, not active. Given that tumors can apparently live with two mutated copies, the idea of further inactivating complexes with ARID1A mutations is probably not a profitable one. But, perhaps there is a ying-yang relationship with another chromatin regulator which can be leveraged. In other words, perhaps inhibiting an opposing complex could restore balance to the cell's chromatin regulation and inhibit the tumor. That's the sort of work which can build off of the foundation these two cancer genomics papers have provided.

Kimberly C. Wiegand, Sohrab P. Shah, Osama M. Al-Agha, Yongjun Zhao, Kane Tse, Thomas Zeng, Janine Senz, Melissa K. McConechy, Michael S. Anglesio, Steve E. Kalloger, Winnie Yang, Alireza Heravi-Moussavi, Ryan Giuliany,Christine Chow, John Fee, Abdalnas (2010). ARID1A Mutations in Endometriosis-Associated Ovarian Carcinomas New England Journal of Medicine : 10.1056/NEJMoa1008433

Jones S, Wang TL, Shih IM, Mao TL, Nakayama K, Roden R, Glas R, Slamon D, Diaz LA Jr, Vogelstein B, Kinzler KW, Velculescu VE, & Papadopoulos N (2010). Frequent Mutations of Chromatin Remodeling Gene ARID1A in Ovarian Clear Cell Carcinoma. Science (New York, N.Y.) PMID: 20826764

Worse Could Be Better

2010-08-31T00:22:00.003-04:00

My eldest brother was in town recently on business & in our many discussions reminded me of the thought-provoking essay "The Rise of 'Worse is Better'". It is on a thought train similar to Clayton Christensen's books -- sometimes really elegant technologies are undermined by ones which are initially far less elegant. In the "WiB" case, the more elegant system is too good for its own good, and never gets off the ground. In Christensen's "disruptive technology" scenarios, the initially inferior serves utterly new markets priced out by the more elegant approaches, but the inferior technology then nibbles slowly but surely to replacing the dominant one. But a key conceptual requirement is to evaluate the new technology on the dimensions of the new markets, not the existing ones.

I'd argue that anyone trying to develop new sequencing technologies would be well advised to ponder these notions, even if they ultimately reject them. The newer and more different the technology, the longer they should ponder. For it is my argument that there are indeed markets to be served other than $1K high quality canid genomes, and some of those offer opportunities. Even existing players should think about this, as there may be interesting trade-offs that might go after totally new markets.

For example, I have an RNA-Seq experiment off at a vendor. In the quoting process, it became pretty clear that about 50% of my costs are going to the sequencing run and the other 50% of costs to library preparation (of course, within both of those are buried various other costs such as facilities & equipment as well as profit, but those aren't broken out). As I've mentioned before, the costs of the sequencing are plummeting but library construction is not on such a steep trend.

So, what if you had a technology that could do away with library construction? Helicos simplified it greatly, but for cDNA still required reverse transcription with some sort of oligo library (oligo-dT, random primers or a carefully picked cocktail to discourage rRNA from getting in). What if you could either get rid of that step, read the sequence during reverse transcription or not even reverse transcribe at all? A fertile imagination could suggest a PacBio-like system with reverse transcriptase immobilized instead of DNA polymerase. Some of the nanopore systems theoretically could read the original RNA directly.

Now, if the cost came down a lot I'd be willing to give up a lot of accuracy. Maybe you couldn't read mutations out or allele-specific transcription, but suppose expression profiles could be had for tens of dollars a sample rather than hundreds? That might be a big market.

Another play might be to trade read length or quality of an existing platform for more reads. For example, Ion Torrent is projected to initially offer ~1M reads of modal length 150 for $500 a pop. For expression profiling, that's not ideal -- you really want many more reads but don't need them so long. Suppose Ion Torrent's next quadrupling of features came at a cost of shorter reads and lower accuracy. For the sequencing market that would be disastrous -- but for expression profiling that might be getting in the ballpark. Perhaps a 16X the initial chip -- but with only 35bp reads -- could help drive adoption of the platform by supplanting microarrays for many profiling experiments.

One last wild idea. The PacBio system has been demonstrated in a fascinating mode they call "strobe sequencing". The gist is that the read length on PacBio is largely limited by photodamage to the polymerase, so letting the polymerase run for a while in the dark enables spacing reads apart by distances known to some statistical limits. There's been noise about this going at least 20K and perhaps much longer. How long? Again, if you're trapped in "how many bases can I generate for cost X", then giving up a lot of features for such long strobe runs might not make sense. But, suppose you really could get 1/100th the number of reads (300)-- but strobed out over 100Kb (with a 150bp island every 10Kb). I.e. get 5X the fragment size by giving up about 99% of the sequence data. 100 such runs would be around $10K -- but would give a 30,000 fragment physical map with markers spaced about every 10Kb (and in runs of 100Kb). For a mammalian genome, even allowing for some loss due to unmappable islands, that would be at least a 500X coverage physical map -- not shabby at all!

Now, I won't claim anyone is going to make a mint off this -- but with serious proposals to sequence 10K vertebrate genomes, such high-throughput physical mapping could be really useful and not a tiny business.

Who has the lead in the $1K genome race?

2010-08-29T21:46:00.002-04:00

A former colleague and friend has asked over on a LinkedIn group for speculation on which sequencing platform will deliver a $1K 30X human genome (reagent cost only). It is somewhat unfortunate that this is the benchmark, given the very real cost of sample prep (not to mention other real costs such as data processing), but it has tended to be the metric of most focus.

Of existing platforms, there are two which are potentially close to this arbimagical goal (that is, a goal which is arbitrary yet has obtained a luster of magic through repetition).
ABI's SOLiD 4 platform can supposedly generate a genome for $6K, though even with pricing from academic core labs I can't actually buy that for less than about $12K (commercial providers will run quite a bit more; they have the twin nasty issues of "equipment amortization" and "solvency" to deal with).
The SOLiD 4 hq upgrade is promised for this fall with a $3K/genome target. Could Life Tech squeeze that out? I'm guessing the answer is yes, as the hq does not use an optimal bead packing. Furthermore, the new paired end reagents will offer 75 bp reads in one direction but only 25 in the other.
I've never understood why a ligation chemistry should have an asymmetry to it (though perhaps it is in the cleavage step), so perhaps there is significant room for improvement there. Of course, those possible 40 cycles are not free, so whether this would help with cost/genome is not obvious (though it would be advantageous for many other reasons). Though, since they can currently get a 30X genome on one slide longer reads would enable packing more genomes per slide & perhaps that's where the accounting ends up favoring longer reads.

Complete Genomics is the other possible player, but we have an even murkier lens on the reagent costs per genome, given that Complete deals only in complete genomes and only in bulk. But, they do have to actually ensure they are not losing money (or at least, with their IPO they won't be able to hide the bleed). Indeed, Kevin Davies (who has a book on $1K genomes coming out) replied on the thread that Complete Genomics has already declared to be at $1K/genome in reagent costs. Perhaps we should move the target to something else (Miss Amanda suggests that $1K canid genomes are far more interesting).

What about Illumina? With HiSeq, they are supposedly at $10K/genome with the HiSeq and many have noted that
the initial HiSeq specs were for a lower cluster packing than many genome centers achieve. That also brings up an interesting issue of consistency -- how variable are cluster packings & therefore the output per run. In other words,
what sigma are we willing to accept in our $1K/genome estimate? Also, the HiSeq specs were for shorter reads than the 2 x 150 paired end
reads that are quite common in 1000 genomes depositions in the SRA (how much longer can Illumina go?).

So, perhaps any of these three existing platforms might meet the mark (454 is a non-starter; piling up data cheaply is not
its sweet spot). What about the ones in the wings? Of course, these are even murkier and we must rely even more on their maker's
projections (and potentially, wishful thinking).

IonTorrent's technology (to be re-branded by Life Tech?) isn't nearly there right now. For $500 (the claim is) you'd get 150Mb of data, or about 0.1X for $1000, so we need about 300X improvement. However, there should be a lot of opportunity to improve. The one touted most in the past is further improvement in the feature density; Ion Torrent was apparently already working on a chip with about 4X the number of features. If we round 300 to 256, then that would only be 4 rounds of quadruplings. If Life could pump those out every 6 months, then that would only be two years to a $1K genome. Who knows how realistic that schedule would be?

But IonTorrent could push on other dimensions as well. Because the flowcell itself is a huge chunk of the cost of a run, squeezing longer read lengths should be possible. Since 454 gets nearly 500 basepair reads routinely (and up to a kilobase when things are really humming), perhaps there is a factor of nearly 4 to get from longer reads. In a similar manner, a paired-end protocol could potentially double the amount of sequence per chip (at a cost of perhaps a bit more than double the runtime; not such a big deal if the run is really an hour). Could that be done? I think I have the schematic for an approach (which might also work on 454); trade proposals for sequencing instruments will be put to my employer for consideration! Finally, as noted in a thread on SEQAnswers, IonTorrent is apparently achieving only about a 1/8th efficiency in converting chip features to sequence-generating sites; better loading schemes might squeeze another few fold out. So perhaps IonTorrent really is 1-2 years away from having $1K genomes (much more likely the 2).

Moving on, could Pacific Biosciences (or the Life tech StarLight (nee VisiGen)) technology have a shot? Lumping them together (since we have virtually no price/performance information for StarLight), PacBio is initially promising $100 runs generating ~60Mb, so $1K would get you about 0.2X coverage, or about 150-fold off, which we'll round to 128-fold or 7 doublings. I think they've already been said to be testing a chip with twice the density, plus a better loading scheme to yield around 2X -- so perhaps it's only 5 doublings.

Finally, there are the technologies which haven't yet demonstrated the ability to read any DNA, but could do so and then move quickly (or not). In this category are any nanopore-based systems (which is a dizzying array of approaches) and Gnu Bio's sequencing-by-synthesis-in-nanodrops approach. And perhaps a few more. These don't even work yet, so even speculative price performance information isn't available.

Finally, a quick note about what a $1K genome means. The X-prize folks have set very strong standards, standards which are far beyond what any short read technology could hope to accomplish and also far beyond what many sequencing applications need. The organizers did not super-design them for no reason; there are applications which need that rigor and also it will greatly cut down on false positives. But, as the regular stream of papers shows, much lower standards will suffice to get interesting biology of whole human genomes.

Lawyers v. Research Funding?

2010-08-24T08:51:00.002-04:00

An ongoing personal quest is to attempt to fill in the gaps in my original education, particularly outside the areas of science in which I feel there exist gaping chasms. Through Wikipedia, books and especially recorded college courses, I slowly patch up what the deficiencies of my education (or all too commonly, my youthful deficiencies in attention during that education) have failed to cover. I'm currently making a third pass through a wonderful course on Roman history since I enjoyed it very much the first two times.

During Rome's early expansion it was ruled by rotating sets of elected officials under a system known to us as the Roman Republic. A series of events (known to scholars as the Roman Revolution) over many decades disrupted this system, culiminating in the replacement of the Republic with the military dictatorship of the Emperors, which would remain until the fall of the empire. An initiating event in the Revolution was an official named Tiberius Gracchus, who in the service of high-minded ideals (rewarding landless soldiers with their own plots on which to support themselves), changed the nature of Roman politics by introducing mob violence to the process (as well as a certain degree of ruthlessness in dealing with the opposition of colleagues).

I fear that yesterday's court decision regarding embryonic stem cell research represents a similar horrible turn. Now, what most commentators will focus on is the very issue of creating human embryonic stem cells and whether the government should finance this. This is an area in which the proponents of both sides of the issue have deeply and sincerely held beliefs which I feel must be respected, though in the end they are fundamentally irreconcilable. But peripheral to that, the case represents a very scary intrusion of lawyers into the research funding process.

One of the claims made by the plaintiffs (in particular, the research James Sherley) is that the new guidelines on what embryonic stem cell research can be funded represent a very real cause of harm to those working on adult stem cell research; they will have more competition for research funding. That is certainly true; if we view research funding for stem cells as a zero sum game (and that is another whole can of balled waxworms I won't dela with). The danger now is that every possible change in federal (or even private?) funding aim will be an opportuntity for litigators to intrude. Wind down project X to fund project Y? LAWSUIT! Either this will dissuade funding from the ebb and flow which is necessary, or a far worse than zero sum game ensues in which funding for science instead funds litigation (or the buy-offs of potential suits which are routine in that field).

Can this genie be stuffed back in the bottle? I'm not legally trained enough to know. Perhaps it was inevitable. Perhaps we need Congress to explicitly forbid it (but would that be legal?) -- and what are the chances of that? Has a terrible Rubicon been crossed; I hope I am wrong in thinking it has.