By Ben Butkus
Pacific Biosciences this week said that it is partnering with several companies, including four that provide target enrichment and sample preparation platforms, to develop and validate protocols for its forthcoming third-generation DNA sequencing platform.
For the partner companies, which include Agilent, Fluidigm, NuGen, and RainDance Technologies, the alliance gives them a head start on grabbing a piece of the sequencing market share that Pacific Biosciences is expected to secure when it launches the Single Molecule Real Time DNA sequencing instrument later this year.
Meantime, Pacific Biosciences is hoping to make the SMRT instrument "technology agnostic" by co-developing protocols for a variety of target enrichment and sample prep platforms, Donna Wilson Earley, product manager at Pacific Biosciences and head of its consumables partnering program, told PCR Insider this week.
"There are a lot of different solutions out there" Earley said. "We want to enable our customers to be able to work with whatever technology they choose."
Although they all compete in the same market, the individual target enrichment and sample prep technologies chosen by Pacific Biosciences "all have certain strengths," Earley said.
Agilent "was an easy pick to partner with," according to Earley, because it has a broad installed base for its SureSelect target enrichment system. "That was our strategy and main reason for picking them," Earley said. "As we begin to build our own installed base for target enrichment protocols and applications, we really want to make sure we're compatible with the majority of the solutions our customers will be using."
Agilent's SureSelect platform is based on hybrid selection and is performed in microcentrifuge tubes or microtiter plates. According to the company, the platform was originally designed to work with Illumina's Genome Analyzer end-sequencing protocol, but is now also optimized to work with Illumina Genome Analyzer paired-end sequencing and Applied Biosystems' (Life Technologies) SOLiD System.
A spokesperson for Agilent told PCR Insider that it was too early in the Pacific Biosciences partnership program to provide additional comment on Agilent's participation.
Meantime, partnering with Fluidigm "provides an intriguing opportunity for [the SMRT] platform," Earley said.
Fluidigm's Access Array system is a microfluidic chip that allows users to amplify 48 specific amplicons from 48 unique samples, in effect preparing 48 libraries in about four hours, according to the company.
"One of the differentiating features of the SMRT platform is the very fast time to result," Earley said. "Our system is able to do sequencing runs in 15 minutes and cycle through many samples in a day." As such, pairing SMRT with Access Array "could be the quickest way to seed our instruments in a high-throughout fashion for targeted sequencing," Earley added. "[Fluidigm] is really paying attention to how to get the largest number of targeted samples out there quickly."
Martin Pieprzyk, a product marketing manager for Fluidigm, told PCR Insider that for the SMRT platform, Fluidigm will likely modify a traditional amplicon-tagging protocol that will allow a user to "basically take 48 different samples and capture or target enrich a specific number of genes from each sample, so the final product coming out of the Access Array is ready to go on the sequencer and doesn't require any additional library preparation."
An added benefit of Fluidigm's platform is a barcoding feature that eliminates the need for traditional library preparation. "This is a six- to eight-position sequence where each sample has a slightly different sequence," Pieprzyk said. "Then you can pool all 48 samples together in post-processing, and sequence them at the same time. This increases your sample throughput anywhere from 48- to 200-fold."
With NuGen, Pacific Biosciences is focusing primarily on its WGA DNA sample-prep system and WTA RNA sample-prep systems, Earley said.
NuGen's Ovation WGA System is a simple linear amplification technology based on single primer isothermal amplification, which replicates genomic DNA from clinical samples for a variety of downstream applications, according to the company.
Meantime, NuGen's Ovation Pico and PicoSL WTA Systems are used to prepare amplified cDNA from a range of 500 picograms to 50 nanograms of total RNA in less than five hours. The kits are specifically designed for applications with very low RNA input requirements for sensitive detection of transcripts over a wide range of abundance, according to the company.
"NuGen has a lot of different solutions, not just for whole-genome or whole-transcript amplification, but target enrichment as well," Earley said. "We are trying to push their [WGA and WTA] kits to the longer and longer read lengths we're going to be requiring in the event our customers don't have enough input sample, or have compromised [formalin-fixed, paraffin-embedded] samples in cancer genetics studies."
Finally, Pacific Biosciences wanted to ensure that its sequencer is compatible with RainDance Technologies' microdroplet-based RDT 1000 system and sequence enrichment kits because RainDance "is an up-and-coming company generating a lot of buzz … and gaining market share rather quickly," Earley said.
"Because of their microdroplet-based technology, and the fact that we're a single-molecule company, they introduce probably the least amount of bias into their system through their approach," Earley said. "We're very keen of taking advantage of that, especially with our sensitive levels. We could go possibly beyond just basic target enrichment applications."
RainDance's Chief Commercial Officer Chris McNary told PCR Insider that RainDance's platform is agnostic to all next-generation sequencing platforms "and we're also working with all third-generation" providers.
RainDance's technology is based on PCR, "which has been identified as the gold standard," McNary said. Its main benefit for target enrichment is that it "can cover 100 percent of the genome, while hybridization can't make that claim," he added. In addition, "the scale we work at, in the nanoliter range, greatly reduces the usage of materials and increases speed," McNary said.
"Pacific Biosciences saw with us a performance advantage and a minimal amount of bias," McNary said. He said that combining RainDance's technology with SMRT may be particularly useful for target enrichment for resequencing applications. Because of its single-molecule capabilities, RainDance's platform is also suited to other certain applications that Pacific Biosciences will be pursuing, McNary said, but declined to elaborate, citing confidentiality agreements.
For each of the partnering companies, the benefit of jumping on the Pacific Biosciences boat seemingly outweighs the fact that their technologies are more or less in direct competition.
Earley said that each of the companies "has been very eager to partner with us. Certainly everyone has questions about whether we are talking with other target-enrichment technology providers. But they are also all aware that each one serves its own specialized niche."
"The biggest benefit to us is market share," Fluidigm's Pieprzyk said. "Right now we work with 454 and Illumina, and obviously with Pacific Biosciences coming on the scene, the market will expand, and being able to work with their instrument will greatly increase our market potential."
Pieprzyk agreed that Pacific Biosciences partnering with multiple companies "has its benefits" because each target-enrichment technology has its strengths and drawbacks.
"The amount of partners may be a little bit of overkill, but you still probably want to have two or three different technologies for target enrichment to cover all of your bases," he said. "If you go from someone who wants to do the whole exome to someone who wants to do a few genes, there currently is no technology that can really do both. There are people who claim they can do both, but you really have to go from hybridization to PCR."