Two startup companies are entering the DNA sample preparation market, and are working on applications for constructing second-generation sequencing libraries.
Boreal Genomics rolled out its first instrument to early-access customers last year. The system uses the company's Synchronous Coefficient of Drag Alteration, or SCODA, technology for electrophoresis-based nucleic acid purification. This week, the company is presenting a second instrument, Aurora, which includes certain improvements, at the Lab Automation meeting in Palm Springs, Calif.
Meanwhile, Sage Science is developing an automated preparative system for electrophoresis-based DNA size fractionation and DNA extraction, called Pippin Prep, which it hopes to launch in about a year.
While Boreal is using rotating electric fields to selectively concentrate nucleic acids in a gel and separate them from contaminants, Sage is aiming to automate the standard lab procedure for preparing nucleic acids from an agarose gel.
One application both companies are pursuing is the purification of size-selected DNA fragments for sequencing libraries for the Roche/454, Illumina, and Applied Biosystems SOLiD sequencers.
Boreal Genomics, based in Vancouver, BC, was founded in 2007 as a spinout from Andre Marziali's lab at the University of British Columbia. The company is based on technology invented by Marziali, an associate professor of biophysics and director of engineering physics at the university, and a colleague. It uses alternating electric fields to move charged molecules in a gel or fluid.
Marziali and his co-workers found that nucleic acids behave differently from other biomolecules in response to these alternating fields, allowing them to focus DNA in the center of an agarose gel, while washing out most contaminants.
They found that their SCODA technology was particularly effective at removing contaminants that other DNA extraction methods had trouble with, for example humic acids from soil samples, which are often used in metagenomic studies.
In a collaborative proof-of-concept study that they plan to publish shortly, Marziali and his collaborators discovered approximately 200 new organisms by extracting and sequencing DNA from tar sand where other DNA-purification methods had failed.
After removing petroleum from the sample, "we threw the sand and everything else directly onto our instrument, and in one step we got DNA out that was clean enough [for cloning and sequencing]," said Marziali, who also serves as Boreal Genomics' president and CSO.
Besides being able to remove contaminants, the SCODA technology can extract DNA from samples that contain DNA down to zeptomolar concentrations, which corresponds to just a handful of DNA molecules per milliliter, he added.
In 2007, Boreal started to build early instrument prototypes and began shipping them to a small number of development partners in late 2007 and 2008.
That instrument, the SCODA 1.3 Development System, is now widely available to customers. It has a list price of $30,000 and advertised consumables costs of 3 cents per run. Users pour their own agarose gels for the instrument, which has a typical run time of about two hours.
"It's meant for people that have challenging samples that are not so concerned about throughput," according to Marziali.
This week, the company is showcasing a new instrument at the Lab Automation meeting that incorporates improvements it has made over the last year or so. That instrument, called Aurora, "has been designed for ease of use," according to the company's website. It uses disposable gel cartridges, and run times have
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been shortened to 5 to 10 minutes. Moreover, it can size-select DNA "so you can do clean-up and length selection in one shot," according to Marziali.
In addition, he and his colleagues recently demonstrated that they can extract nucleic acids based on their sequence, with resolution down to a single nucleotide difference between two samples. Unlike microarrays, this method is not prone to non-specific hybridization since it uses electric forces, according to Marziali.
Boreal is currently looking for a small number of users for an early-access program for Aurora and plans to distribute the instrument more broadly six to 12 months from now. The company has yet to determine the price for the instrument after performing "cost-reduction engineering," he said.
Eventually, Aurora will replace the old instrument, but the company also hopes that it will open new applications beyond soil metagenomics. For example, the technology could be used for pathogen and biomarker detection in body fluids or in food safety testing, where current testing methods are expensive and slow, Marziali said.
"This is a very broad platform technology; we are able to get into markets that are anywhere from homeland security and forensics to food safety and clinical diagnostics, so we have a number of irons in the fire, so to speak, and are exploring those opportunities individually," Marziali said.
For second-generation sequencing, the main application will be in library construction, he said. "We are pursuing length selection methods that will allow you to extract DNA … all that in one shot," he said. "We can already do length selection; the question is, will we be able to do it over a tight enough range that would be appropriate for that kind of sequencing?"
Ken Dewar, an associate professor in the department of human genetics at McGill University, has had a SCODA 1.3 Development system since last fall and has used it to extract DNA from soil for 454 sequencing projects. He plans to use the system also for size selection, either for 454 sequencing libraries or for large-insert cloning.
The machine has been "very straightforward to operate" and requires no proprietary reagents, according to Dewar. One of the challenges, he said, is that the output DNA from a single SCODA run is currently insufficient for a 454 library, but he and his colleagues are working on improving the SCODA yield as well as reducing the DNA input requirements for 454.
"Where we see the importance of the SCODA is in places where there is no prep and where [people] have tried the conventional preps and they just don't work," he said. "Then the SCODA would be our backup plan for that."
Meantime, Marziali and his colleagues are working on further improvements to the technology, both in his lab at UBC and at the company, which holds an exclusive license to the technology and licenses new developments from UBC. The company's board chairman is Tom Willis, a founder and former CSO of ParAllele BioScience whom Marziali knows from graduate school.
About eight scientists are working on the SCODA technology, about half of them in Marziali's lab under grant funding and half at Boreal Genomics, which recently moved into its own space in an incubator building near the university.
So far, the company has grown "as non-dilutively as possible," according to Marziali. This month, Boreal raised an undisclosed amount of funding in a first seed financing round, mostly from angel investors. It has also raised approximately $1.5 million in grant funding, including an award last year from the US National Human Genome Research Institute to develop the technology further for metagenomic applications.
Also last year, Marziali's lab won a grant from the US National Cancer Institute to enable the SCODA technology to concentrate tumor DNA in body fluids.
Boreal Genomics also received "a fair amount of capital" from sales of its early instrument to early access users, which helped to fund part of the development of Aurora.
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Depending on its strategy — whether it seeks a distribution partner or opts to build its own sales channels — Boreal Genomics might require more funding in the future, according to Marziali. However, "we are not looking to go out for fundraising immediately," he said.
Meantime, Sage Science is working on an automated system for preparative agarose gel electrophoresis and extraction of DNA, called Pippin Prep, which it initially plans to market for size fractionation of sheared DNA for second-generation sequencing libraries.
The Beverly, Mass.-based company aims to "develop instruments and consumables to simplify processes that molecular biologists do every day," according to Chris Boles, its vice president and chief scientific officer.
As Sage Science was considering agarose gel electrophoresis in general, "during our initial market research, we came across some next-generation sequencing laboratories, and they were using tremendous numbers of agarose gels to prepare their library samples for sequences," Boles recalled.
"It's a fundamental step in the common next-generation sequencing protocols" for systems that use amplified DNA such as Illumina's Genome Analyzer, Roche's 454 GS FLX, and Applied Biosystems' SOLiD, and size selection is especially important for paired-end libraries, he said. The process is currently very labor-intensive, requiring users to run DNA on an agarose gel, cut out the desired DNA fragments, and extract the DNA from the gel.
Sage Science's goal is to cut the labor associated with the process by up to 90 percent. Its system will use pre-cast agarose gel cassettes that will be processed in an instrument with an optical reading station and an electrode system that determines the size of the DNA. The system then extracts the DNA into a liquid-filled elution chamber.
So far, the company has developed prototypes that are "are working quite well," Boles said, adding that the company hopes to launch an instrument about a year from now. Boles is scheduled to give a talk about the technology at the Advances in Genome Biology and Technology conference in Marco Island, Fla., next week.
Sage was founded in 2005 by Gary Magnant, a founder of electrophoresis equipment manufacturer Owl Scientific, and spent its first two years developing two OEM products for other companies.
The automated DNA-selection and -extraction system, which Sage has been developing since last spring, will be the first product that the company will sell under its own name.