New method for "bottom-up genome assembly"

Itaya, et al., have published a new method for assembling ~5kB DNA fragments into genome-sized pieces in this month's Nature Methods (PubMed).  Jason Kelly has launched a blog, Free Genes, where he describes the new method.  Welcome to the blogosphere, Jason.

I won't add anything to Jason's post, other than to note that because Itaya's method exploits a recombination mechanism present in a microbe, there is no need to manipulate large pieces of DNA "by hand".  This is a significant advantage over methods that require lots of pipetting between PCR steps, which exposes the growing DNA to fluid shear.  The reliance upon natural mechanisms for assembly might mean the method is better suited to the garage than something that uses fluid transfer.

Finally, building ~5kB segments doesn't appear to be such a big deal at this point.  While Itaya's method isn't completely general, and as described may be a bit slow, it should be widely useful to anyone who has an in-house method for making gene-sized pieces of DNA and who doesn't want to pay a foundry to assembly even larger pieces.

(Update: Oops.  I forgot to add that this sort of thing is just what I suggested in my previous post, when I observed that while Venter may have made excellent progress in building an artificial chromosome he certainly doesn't have a lock on building new organisms.)

Updated "Longest Synthetic DNA" Plot

With the reported completion of a 580 kB piece of DNA by Venter and colleagues, it is time to update another metric of progress in biological technologies.  Assuming the report is true, it provides evidence that the technological ability to assemble large pieces of DNA from the short oligonucleotides produced by DNA synthesizers is keeping up with the productivity enhancements enabled by those synthesizers (see my prior post "Updated, Um, Carlson Curve for DNA Synthesis Productivity").  That said, this is an accomplishment of art and science, not of commerce and engineering.  The methods are esoteric and neither widespread nor sufficiently low cost to become widespread.

The news report itself is a couple of months old now.  It yet to be confirmed by scientific publication of results, so I am breaking my habit of waiting until I can see the details of the paper before including another point on the plot.  Perhaps I just need something to do as a break from writing my book.

In any event, in the 6 October, 2007 edition of The Guardian, Ed Pilkington reported, "I am creating artificial life, declares US gene pioneer":

The Guardian can reveal that a team of 20 top scientists assembled by Mr Venter, led by the Nobel laureate Hamilton Smith, has already constructed a synthetic chromosome, a feat of virtuoso bio-engineering never previously achieved. Using lab-made chemicals, they have painstakingly stitched together a chromosome that is 381 genes long and contains 580,000 base pairs of genetic code.

It does not appear, from Mr. Pilkington's story, that Venter et al have yet inserted this mammoth piece of DNA into a cell.  Though Craig Venter is supposedly "100% confident" they can accomplish this, and as a result will boot up a wholly artificial genome running a semi-artificial organism; "The new life form will depend for its ability to replicate itself and metabolise on the molecular machinery of the cell into which it has been injected, and in that sense it will not be a wholly synthetic life form."

The Guardian story includes a comment from the dependably well-spoken Pat Mooney, director of the ETC Group.  Says Mooney,  "Governments, and society in general, is way behind the ball. This is a wake-up call - what does it mean to create new life forms in a test-tube?"

Here is an open letter to Mr. Mooney:

Dear Pat,

It doesn't mean a damn thing.  Except that it helps you raise more money by scaring more people unnecessarily, so that you can go on to scare yet more people.  Have fun with that.

Best Regards,

Rob Carlson

PS Great business model. 

I just can't get really excited about 580 kB of synthetic DNA.  First, while interesting technically, the result is entirely expected.  People keep saying to me that it is really hard to manipulate large pieces of DNA in the lab, and to this I say many things we do are really hard.  Besides, nature has been manipulating large pieces of DNA very successfully for a while now.  Say, three billion years, give or take.  It was inevitable we would learn how to do it. 

Second, I know of a few individuals who are concerned that, because there is insufficient funding for this sort of work, Venter and his crew will now have some sort of lock on the IP for building new organisms.  But it is so very early in this technological game that putting money on the first demonstrated methodology is just silly.  Someone else, probably many different someones, will soon demonstrate alternatives.  Besides, how many times are we going to need to assemble 580,000 bases and 381 genes from scratch?  The capability isn't really that useful, and I don't see that it will become useful anytime soon.

The more interesting numbers are, say, 10-50 genes and 10,00-50,000 bases.  This is the size of a genetic program or circuit that will have interesting economic value for many decades to come.  But while assembling synthetic constructs (plasmids) this size is still not trivial, it is definitely old news.  The question is how will the cost for constructs of this size fall, and when can I have that DNA in days or hours instead of weeks?  And how soon before I can have a desktop box that prints synthetic DNA of this length?  As I have previously noted in this space, there is clear demand for this sort of box, which means that it will happen sooner or later.  Probably sooner. 

Third, the philosophical implications of constructing an artificial genome are overblown, in my humble opinion.   It is interesting to see that it works, to be sure.  But the notion that this demonstrates a blow against vitalism, or against other religious conceptions of life is, for me, just overexcitement.  Venter and crew have managed to chemically synthesize a long polymer, a polymer biologically indistinguishable from naturally occurring DNA; so what?  If that polymer runs a cell the same way natural DNA does, as we already knew that it would, so what?  Over the last several millennia religious doctrine has shown itself to be an extremely flexible meme, accommodating dramatic changes in human understanding of natural phenomena.  The earth is flat!  Oh, wait, no problem.  The earth is at the center of the universe!  No?  Okay, we can deal with that.  Evolution is just another Theory!  Bacteria evolve to escape antibiotics?  Okay, God's will.  No problem. I can't imagine it will be any different this time around.

Finally, it is worth asking what, if any, implications there are for the regulatory environment.  The Guardian suggests, "Mr Venter believes designer genomes have enormous positive potential if properly regulated."  This is interesting, especially given Venter's comments last winter at the initial public discussion of "Synthetic Genomics: Options for Governance".  I don't know if his comments are on record anywhere, or whether my own public comments are for that matter, but Venter basically said "Good luck with regulation," and "Fear is no basis for public policy."  In this context, I think it is interesting that Venter is not among the authors of the report.

I just finished writing my own response to "Options for Governance" for my book.  I can't say I am enthusiastic about the authors' conclusions.  The  authors purport to only present "options".  But because they examine only additional regulation, and do not examine the the policy or economic implications of maintaining the status quo, they in effect recommend regulation.  One of the authors responded to my concerns of the implicit recommendation of regulation with, "This was an oversight."  Pretty damn big oversight.

Today's news provides yet another example of the futility of regulating technologies to putatively improve security.  Despite all the economic sanctions against Iran, despite export restrictions on computer hardware, scientists and engineers in Iran report that they have constructed a modest supercomputer using electronic components sold by AMD.  Here is the story at ITNews (originally via Slashdot).  Okay, so the Iranians only have the ability to run relatively simple weather forecasting software, and it may (may!) be true that export restrictions have kept them from assembling more sophisticated, faster supercomputers. (I have to ask at this point, why would they bother?  They are rolling in dollars.  Why not just pay somebody who has a faster machine to do the weather forecasting for you?  It suggests to me that they have pulled the curtain not from their best machine, but rather from one used to be used for weapons design and is now gathering dust because they have already built a faster one.)  Extending this security model to biological technologies will be even less successful.

Export restrictions for biological components are already completely full of holes, as anyone who has applied for an account at a company selling reagents will know.  Step 1: Get a business license.  Step 2: Apply for account.  Step 3: Receive reagents in mail.  (If you are in a hurry, skip Step 1; there is always someone who doesn't bother to ask for it anyway.)  This particular security measure is just laughable, and all the more so because any attempt to really enforce the legal restrictions on reselling or shipping reagents would involve intrusive and insanely expensive physical measures that would also completely crimp legitimate domestic sales.  I can only imagine that the Iranians exploited a similar loophole to get their AMD processors, and whatever other hardware they needed.

Well, enough of that.  I have one more chapter to write before I send the book off to reviewers.  Best get to it.

Updated, um, Carlson Curve for DNA Synthesis Productivity

It seems that productivity improvements in DNA synthesis have resumed their previous pace.  As I noted in Bio-era's Genome Synthesis and Design Futures, starting in about 2002 there was a pause in productivity improvements enabled by commercially available instruments.

According to the specs and the company reps I met at iGEM 2007, a single Febit "Geniom" synthesizer can crank out about 500,000 bases a day and requires about 30 minutes of labor per run.  It looked to me like the number should be closer to 250KB per instrument per day, so I made an executive decision and allowed that the 16 synthesizers one person could run in a day could produce 2.5 megabases of single-stranded ~40-mers per day.  This in part because there is some question about the quality of the sequences produced by the particular chemistry used in the instrument.  It was asserted by the company reps that the Geniom instruments are being adopted by major gene synthesis companies as their primary source of oligos.  Note that running all those instruments would cost you up front just under US$ 5 million, without volume discounts, for 16 of the $300,000 instruments (plus some amount for infrastructure).

The quality of the DNA becomes particularly important if you are using the single-stranded oligos produced by the synthesizer to assemble a gene length construct.  To reiterate the point, the 2.5 megabases per day consists of short, single-stranded pieces.  The cost -- labor, time, and monetary -- of assembling genes is another matter entirely.  These costs are not really possible to estimate based on publicly available information, as this sort of thing is treated as secret by firms in the synthesis business.  Given that finished genes cost about 10 times as much as oligos, and that synthesis firms are probably making a decent margin on their product, the assembly process might run 5 to 8 times the cost of the oligos, but that is totally a guess.  (Here is a link to a ZIP file containing some of the graphics from the Bio-era report, including cost curves for gene and oligo synthesis.)

One final note: the Febit reps suggested they are selling instruments in part based on IP concerns of customers.  That is, a number of their customers are sufficiently concerned about releasing designs for expression chips and oligo sets -- even to contract manufacturers under confidentiality agreements -- that they are forking over $300,000 per instrument to maintain their IP security.  This is something I predicted in Genome Synthesis and Design Futures, though frankly I am surprised it is already happening.  Now we just have to wait for the first gene synthesis machine to show up on the market.  That will really change things. 

The Moon in HD

Super.  Duper.  Cool.

How big is the Bio-economy?

The words "biotechnology" and "biotech" are often used by the press and industry observers in limited and inconsistent ways.  Those words may be used to describe only pharmaceutical products, or in another context only the industry surrounding genetically modified plants, while in yet another context a combination of biofuels, plastics, chemicals, and plant extracts.  The total economic value of biotechnology companies is therefore difficult to assess, and it is challenging to disentangle the component of revenue due each to public and private firms.

I've managed to get a rough idea of where the money is for industrial biotech, agbiotech, and biopharmeceuticals.  Based on surveys from Nature Biotechnology, the U.S. Government, various organizations in Europe, and several private consulting firms, it appears estimates of total revenues range from US$ 80 to 150 billion annually, where the specific dollar value depends strongly on which set of products are included.  The various surveys that provide this information differ not only in their classification of companies, but also in methodology, which in the case of data summarized by private consulting firms is not always available for scrutiny.  For whatever reason, these firms tend to produce the highest estimates of total revenues.  Further complicating the situation is that results from private biotech companies are self-reported and there are no publicly available documents that can be used for independent verification.  One estimate from Nature Biotechnology, based on data from 2004 (explicitly excluding agricultural, industrial, and environmental biotech firms), suggested approximately 85% of all biotech companies are private, accounting for a bit less than 50% of employment in the sector  and 27% of revenues.

A rough summary follows:  As of 2006, biotech drugs accounted for about US$ 65 billion in sales worldwide, with about 85% of that in the U.S.  Genetically modified crops accounted for another US$ 6 billion, with industrial applications (including fuels, chemicals, materials, reagents, and services) contributing US$ 50-80 billion, depending on who is counting and how.  Annual growth rates over the last decade appear to be 15-20% for medical and industrial applications, and 10% for agricultural applications.

I am not going to go through all the details here at this time.  But the final amount is pretty interesting.  After sifting through many different sets of numbers, I estimate that revenues within the US are presently about US$125 billion, or approximately 1% of US GDP, and growing at a rate of 15-20% annually.

1% of GDP may not seem very large, but a few years ago it was only 0.5%.  At some point this torrid growth will have to slow down, but it isn't clear that this will be anytime soon.  Nor is it clear how large a fraction of GDP that biotech could ultimately be.  That is my next project.

Off to iGEM 2007

I am headed out the door to the 2007 International Genetically Engineered Machines (iGEM) Competition at MIT.  There look to be ~56 teams composed of ~400 students from around the world.  As I am a judge this year, I won't be blogging any more about it until it's over.

I have been looking forward to this for months -- it should be great fun.

More Skyscraper Tourism

I'm in Houston today, speaking to a small group of American Petroleum Institute members about the future of biofuels  (Thanks for the recommendation, Jamais).  More air travel in the service of explaining carbon neutral fuels.  Nonetheless, the view from the 49th floor of the Shell Plaza tower is remarkable.

I am told by residents that Houston has evidently just decided the answer to local traffic and transit issues is to widen I-10 from 15 to 22 lanes.  They are apparently proud of the 2 HOV lanes that will come with the expansion.

Huh.

Oil on the way to a C-note per barrel

Over the last several months I have had the opportunity to talk to a good number of oil and gas executives.  Way back, say, just last summer,  when oil was only at $65 per barrel, it still seemed like a stretch for the oil guys (yes, all guys) to claim oil would hit $100 by the end of 2007.  Now, according to Reuters, oil has just hit $92 per barrel, up 30% since August alone.  It seems the C-note per barrel may arrive quite soon.

Part of this run up in price is due to the decline of the dollar, which may further encourage a shift to trading oil in euros.  The follow on effect of moving so much trade away from the dollar can't be seen as a good sign for the U.S. economy.  Yes, it is still true that foreign governments still hold large amounts of U.S. government debt, which will always be denominated in dollar and which gives many nations an interest in propping up U.S. currency.  But slippage in international use of the dollar for trade makes me even less interested in keeping any business I do here in the U.S.  I have just accepted my first project from Canada, and the contract stipulates pay in Canadian currency.  I might even ponder leaving it in that currency, as the Canadian dollar has been appreciating with respect to the U.S. dollar at a fairly decent rate.

Anyway -- back to oil -- a significant motivation in the increase in price is limitation of supply and increased demand.  Some of this will get fixed as new refining capacity comes on line.  Shell, for instance, is investing US$ 7 billion in doubling the capacity of a refinery in Texas, though this will take many years to come on line.

Bio-era's numbers suggest that by 2020, given current plans, biofuels will amount to 10% of global liquid fuel use.  That doesn't seem like much, but the increase from ~2% to 10% of use will account for 50% of the global increase in use, which is a big deal.  I am beginning to wonder if this is already an underestimate.  Many companies are making good progress in producing various liquid fuels using microbes (see previous posts here and here), and any shift away from the dollar in trading oil will cause further substitution within the U.S.

It will be interesting to see what effect this has on the economics of distributed fuel production.

When microbes innovate faster than humans do

CNN is reporting that  methicillin-resistant Staphylococcus aureus (MRSA) is afflicting a number of high school students in the U.S.  One student has died from an infection apparently contracted at school, while another 15 or so students in two states have tested positive.

This is getting press in part because of a report out in JAMA that the rate of infection from MRSA around the U.S. could be twice as high as previously thought, with a mortality rate of almost 20%.  (Here is the paper on PubMed: "Invasive methicillin-resistant Staphylococcus aureus infections in the United States".)  MRSA was first observed in the U.S. only in 1981.  Thus over only about 25 years we have produced a bug, through profligate use of antibiotics and poor sanitation, that may be a bigger killer than even HIV.

This while NIH funding has more than doubled, where most of that money has gone to established investigators (See my post, "The Death of Innovation, or How the NIH is Undermining Its Future") doing whatever it is they do that doesn't result in new antibiotics.  Where is the Health in NIH?

I heard yesterday via the grapevine that an NIH review panel failed to award any of 19 worthy new grants to younger investigators because all the money in the program is sopped up by existing grants.  You could argue that we should just increase the NIH budget, to which I would be sympathetic, but it is by no means clear that the present funding is well-spent.

The Institute Para Limes

I spent part of last week at the "opening congress" of the Institute Para Limes (IPL) in The Netherlands.  The IPL is meant to be a European version of the Santa Fe Institute (SFI) for the new century, though because of it's cultural mileau it is also meant to be something different.  The meeting last week was supposed to help sort out the focus and style of the place. 

Wikipedia notes that;

SFI's original mission was to disseminate the notion of a separate interdisciplinary research area, complexity theory referred to at SFI as "complexity science". Recently it has announced that its original mission to develop and disseminate a general theory of complexity has been realized. It noted that numerous complexity institutes and departments have sprung up around the world.

SFI was founded by a bunch of famous people, a Nobel Laureate included, and has been much lauded in the press, though its reputation is not universally sterling in academic circles.  This is primarily because, I suspect, many people are still trying to figure out exactly what "Complexity Science" really is all about.  It's a fair question.  But there has been a great deal of good work done at the SFI.

The director of SFI, Geoff West, was the first speaker at the Institute Para Limes meeting, and his talk focussed both on how SFI has succeeded and also his own contributions in the areas of allometric scaling.  He also spoke about this really cool paper in PNAS that I printed out last spring, but have somehow managed to not yet read, "Growth, innovation, scaling, and the pace of life in cities". 

The IPL will eventually be sited in a renovated monastery in Duisberg, which is intellectually, by design, approximately in the middle of nowhere.  This part of the plan for IPL confuses me a bit.  It will take at least 90 minutes to get to IPL from Amsterdam, probably more if you have to change trains multiple times, like I did, and then find a taxi for the final leg.  There is something to be said for making sure you have some intellectual distance from staid Universities, but in my experience it a block or two is usually enough to serve as infinitely high barriers between academic departments.  At Princeton, for many years, it was an exceptionally rare sight for anyone to even cross the street between Jadwin (Physics) and Lewis Thomas (Molecular Biology) for the purposes of a scientific discussion.

The meeting was a chance for me to catch up with Sydney Brenner a bit, to stand by has he and Gerard t'Hooft got into an animated, um, communication, about the purpose of DNA, and to hear Sydney drop a few bon mots:

On "factory science" in biology: "Low input, high throughput, no output."

On evolution: "Mathematics is the art of the perfect.  Physics is the art of the optimal.  Biology is the art of the satisfactory.  Patch it up with sticky tape, tie it up with twine, and go on.  If it doesn't work, end of story, next genome."

Gerard t'Hooft had this nice bit about the process of science: "Science is about the truth.  Science zooms in on the truth.  The truth changes, in part due to changes in science, but the assumptions and conjectures are always periodically tested."

And Science Always Wins.