This article was originally published Jan. 10.
Nabsys is planning to launch a first iteration of its semiconductor-based sequencing system in the second half of the year, company officials told In Sequence last week. The first version will focus on genome mapping of microbial organisms with future versions having applications in assembly and validation of larger genomes, structural variation detection, targeted sequencing, and eventually whole-genome sequencing.
For each application, the system will have a sample-to-result turnaround time of less than a day, Stan Rose, the company's chief commercial officer, told IS.
The system runs DNA fragments with hybridized probes through microfluidic nanochannels on a silicon chip using a semiconductor to read electrical signals elicited from the probes.
Rose calls the technology "positional sequencing," because the probes are used to position fragments of DNA into their correct location on the genome (IS 1/17/2012).
The first iteration of the system will employ a limited number of probes, likely between five and eight, and its primary function will be genome mapping in microbial genomes. In the future, as the company increases the number of probes and the complexity of the probe mixture, the system can be used for whole-genome sequencing, Rose said.
Similar to Life Technologies' Ion Torrent platforms, the basic box will stay the same, while upgrades will involve new chips.
The box will be priced around $50,000 with the chip and reagents at $150 each.
"The goal is to advance the technology so that when we launch it, the cost is less than $1,000 per sample with results delivered in less than a day," Rose said.
Rose said that the company is already working with a handful of early-access users on the system.
Turnaround time will vary depending on the application. Currently, sample prep is around four hours, but Rose said this could be reduced to between one and two hours. The time for running the sample through the chip and collecting data will also vary depending on the application, but the initial application of genome mapping will take no more than an hour. Data analysis and assembly will range between a few seconds for microbial genomes to an hour or so for more complex genomes, Rose said.
After the user extracts DNA using any method, the second step in using the system is probe hybridization. Depending on the application, the number and nature of the probes will vary. For instance, to do genome mapping, a probe master mix will be aliquoted into a well with the DNA sample. But for sequencing, the sample will need to be distributed among many different wells, each with pre-aliquoted probe pools, Rose explained. However, in both cases, the process will be automated and involve minimal hands-on time.
Next, DNA is incubated with a proprietary reagent mix — the "special sauce," Rose said. From the user perspective, all this step requires is pipetting the hybridized DNA into a master mix of reagents, Rose said. Even for the sequencing application, this master mix of reagents will be the same for all the pools.
The probe hybridization and incubation steps take several hours, but Rose said the company is working on reducing that to about one hour.
The DNA is then loaded onto the chip, where it is driven through detectors by a combination of electrophoretic and hydrodynamic forces at a speed of one million bases per second per detector. One important distinction between this approach and those of nanopore sequencers, said Rose, is that in Nabsys' method the goal is not to detect every single nucleotide, but rather to detect where probes are bound to the DNA. This enables the system to move rapidly without giving up accuracy.
The system creates a map of where probes are bound, and by "increasing throughput and probe library complexity, we can create enough maps of fragments so the collective dataset represents the entire genome many times over with independent measurements of the same nucleotide, with different probes of different design and all reassembled," he said, noting that this technique eliminates systematic errors.
For simpler applications like mapping of small genomes, a small number of probes can be used, Rose said. For example, a single 6-mer will bind on average every 4,096 bases. Using four different 6-mers will yield a binding event every 1 kilobase, he said, which is a "pretty high density map" that can be generated from a single reaction in a single tube, with data delivered directly to a laptop in the same day.
For whole-genome sequencing, company researchers have been working with several thousand probes of varying lengths, and working on a 384-well pool creating that many different probe mixtures.
The company has not yet settled on a specified number of probes or probe length for its initial launch, but Rose said the vast majority of probes would be 6- to 8-mers, and that the initial system would likely include fewer than five probes.
In terms of read length, the company has been experimenting with DNA fragments of up to 165 kilobases.
"We'd like the fragments to be as long as possible, but without having the user resort to any DNA isolation techniques not commonly used prior to next-gen sequencing," Rose said.
He added that the company has also tested DNA prep kits from Qiagen, which give a range of fragments, most of which fall in the 30- to 70-kilobase range, "which is very nice for our approach," he said.
Following the initial application of genome mapping, Rose said that there is a big market in cytogenetics — in both the prenatal diagnostic and cancer markets — for which the system would be optimal. This would involve essentially using the system to do "electronic karyotyping," as a noninvasive test to screen for fetal aneuploidy.
Detecting structural variations in cancer will be another important application, he said. For example, a probe mixture could be designed that would detect translocations in a cancer patient and could also screen for specific SNPs associated with drug sensitivity or toxicity, he said.
Aside from clinical applications, the system will have applications in genome assembly and finishing, helping to correct de novo short-read assemblies; and in applied markets for things like pathogen detection, environmental monitoring, and food quality assurance, he said.