After a library is properly prepared, (remember it can be from many sources – randomly sheared genomic DNA, cDNA from a small RNA sample, an immunoprecipitated sample) the library molecules need to be amplified in some manner, before the sequencing takes place. Thus there is a critical need for accurate quantitation of the library DNA, whose importance can be overlooked.
Looking at sequencing from one perspective, library preparation is straightforward. Sequencing a genome (whether bacterial on the order of 5 million bases or a human at 3 billion bases) is a shotgun-based affair with tens of millions to tens of billions of reads that overlap multiple times across the genome (known as ‘fold coverage’). (Thus a 30x human genome coverage would require some 90 billion bases, or a 15x coverage of each haploid allele.) Multiple random start points, 30-fold coverage across the entire genomic sample, one takes a gDNA sample, randomly shears it, attaches synthetic adapters, and off you go following the manufacturer’s protocol on getting sequence data out, whether by Roche / 454, Illumina GAIIx or HiSeq 2000, Life Technologies SOLiD or 5500xl, Pacific BioSciences RS, Illumina MiSeq, Life Technologies Ion Torrent PGM…
As I mentioned in my prior post, Sanger capillary sequencing is not going away anytime soon. Yet next-generation sequencing has made a huge mark in the world – growing from zero in 2005 to a USD $1 Billion market in 2012. And its growth is estimated by various sources to grow 20 to 25% every year for the next five years, approximately tripling in size from where we are now.
Even though the history of next-generation sequencing is short (the 454 GS20 came out in 2005, the Solexa 1G in 2007, and the SOLiD 2 in 2008), there is a robust genomic revolution going on, and a fierce battle in the marketplace with plummeting costs and soaring throughput. Whether Moore’s Law is beat by some 2.5-fold or even faster, there is no question that we are in the middle of burgeoning growth, remarkable discovery, and new insights and discoveries just about every day.
We’re on the verge of a new era in education. This week, the Smithsonian Museum announced a new genomics exhibit at the National History Museum, in conjunction with the National Human Genome Research Institute (NHGRI), and generous grants from several companies and institutions, including a major contribution from Life Technologies (on the order of $3M). The Museum of Natural History already has a nice website reviewing progress in genomics, but this is certainly a welcome development. To quote Eric Green, “It is not enough [for] researchers like me and others at NHGRI to be excited about the human genome and the opportunities before us… because genomic knowledge will become increasingly relevent it is all the more urgent for the public to understand and to appreciate the applications it has for society and for individuals and medical care.”
An enterprising salesperson from Complete Genomics used this newfangled social media thing called LinkedIn to make her mark on the world (perhaps) by posing a discussion question. (It was over at the ‘Genome Interpretation‘ group in case you were wondering.) Entitled, “The last days of exome sequencing“, she posed the question whether exome sequencing day’s were numbered.
We’re living through a time this year of market transition. The Illumina HiSeq continues to have a strong market position (although the upcoming Ion Torrent Proton sets out to change that, however it won’t be available until this September / October). For the time being, customers with NGS platforms are considering upgrading (from the HiSeq 2000 to a ‘fast-mode’ 2500, or from a 5500 to a 5500 ‘Wildfire’ that will be launched later this year, decreasing the cost-per-base substantially and eliminating ePCR).
Yesterday was a good day. Baylor and Life Technologies announce that the upcoming Ion Torrent Proton sequencer has been installed at the Baylor College of Medicine’s Human Genome Sequencing Center. (For those of you who may have missed it, this was announced at the Consumer Electronics Show in Las Vegas in January, and was considered by some ‘today’s coolest gadget‘ there.) Promising a $1,000 genome in 4 hours, a key milestone was met: getting the first ‘early access’ systems into the hands of customers.
In the roads I travel (and it is now over six years since I made that fateful move from being an ‘internal’ Product Manager to ‘customer-facing’ sales representative) the buying process is all about perceptions of the customer. Right or wrong, potential customers each receive the information from a local representative (from whichever vendor) and filter it through their own set of criteria. Opinions they read in their journals of choice, opinions from their valued collaborators and other friends in the research world, tidbits gathered from their post-docs, all form a perception in their mind about what a particular product’s value is to them.
At ABRF last weekend (Association for Biomolecular Resource Facilities) not only was I able to attend George Church’s talk, but I also was able to hear Steven Roever talk about his new nanopore sequencing company, Genia, where he is CEO.
As George Church introduced him, he has an unusual background for a next-generation sequencing company – involved with encryption and Digital Rights Management in prior roles, he met Roger Chien, a Maxim semiconductor company Digital to Analog Conversion chip expert, while at a Sand Hill Road Starbucks, as Roger was reading a book about the origin of life. (I’m not sure exactly which book, but it could have been Freeman Dyson’s.) Maxim, by the way, is a 9500-strong, $2.5B revenue company.
One of the great privileges of working on the marketing side of a life sciences company (compared to the sales side) is the ability to go to a lot of conferences. As a sales representative, it was a hit-or-miss affair, whether I’d be one of the ‘chosen ones’ to go to AGBT, ASHG (now called ICHG), or ABRF.
(For those who are not in genomics, AGBT = Advances in Genome Biology and Technology; ASHG = American Society for Human Genetics; ABRF = Association of Biomolecular Resource Facilities.)
And for a sales rep, it’s a difficult task; you encounter potential customers from many quarters, increasing the sales footprint for the company as a whole, but not so rewarding for your own particular territory. On top of that, you can’t participate in the science per-se, but certainly will have other opportunities to interact with the conference attendees, and glean some of the ‘take-home’ lessons second-hand, if they are so inclined to ask.
Any healthy diet has a good balance: regular ‘staples’, a fair amount of variety, and the occassional surprise ‘treat’.
For next-generation sequencing, the web offers unparalled information to keep up-to-date. Here is a short list of four sites that compose a healthy NGS diet – everyday staples and a lot of variety!