This article was originally published Oct. 27.
NEW YORK (GenomeWeb) – Startup ElectroSeq is looking for funding to develop a highly sensitive semiconductor detection technology, developed at the University of New Mexico, for DNA sequencing. The ultimate goal is to manufacture a sequencing chip with 10 billion wells that would enable researchers to sequence a human genome for $100.
The company's core technology relies on double-gated ion-sensitive field effect transistors, or ISFETs, and was developed and patented by three UNM scientists: Payman Zarkesh-Ha, an associate professor of electrical and computer engineering; Jeremy Edwards, an associate professor in the department of chemical and nuclear engineering; and Steven Brueck, a professor of electrical and computer engineering.
Edwards, a former postdoc in George Church's group at Harvard Medical School, is no stranger to sequencing technology development: several years ago, he was involved in the development of the Polonator, a low-cost sequencing platform out of Church's lab.
According to Paul Szauter, ElectroSeq's founder and CEO, double-gated ISFETs allow photons or hydrogen ions to be measured with greater sensitivity than existing technology, including the one used by Thermo Fisher Scientific's Ion Torrent. While another company is exploiting double-gated ISFETs to boost optical signals in fiber-optic cables, ElectroSeq is planning to put it to use for pH-based DNA sequencing.
Because of the greater sensitivity, fewer copies of template DNA are needed for sequencing, so rolling circle amplification to create so-called "rolonies" could be used instead of bead-based emulsion PCR. Rolonies consist of only 5,000 copies of DNA and are about 300 nanometers in size, much smaller than beads with amplified DNA, Szauter said, and ElectroSeq has a patent pending on the use of rolonies.
The main advantage of using rolonies and double-gated ISFETs, which he said do not require a reference transistor next to each measuring transistor, is the ability to build sequencing chips with higher well densities.
Another advantage is that rolling circle amplification likely has different biases than either emulsion PCR, which is used by Ion Torrent, or bridge amplification, which is employed by Ilumina. This might enable ElectroSeq to sequence parts of the human genome that are currently missed by rival platforms, he said.
In proof-of-concept experiments, Zarkesh-Ha and Edwards have already shown that they can use a prototype chip with 35,000 sensors to detect the incorporation of individual bases into rolony DNA. "It was noisy signal after a few bases, but it was signal that said it's going to work," Szauter said.
Also, at the IEEE International System-on-Chip conference in Las Vegas last month, the researchers presented the concept of using highly sensitive pH-to-current conversion ISFET sensing technology for real-time DNA sequencing, and showed that they were able to measure very small pH changes.
Another paper that has been submitted but not yet published will demonstrate that the sensitivity can be increased tenfold by operating the chip in so-called avalanche mode, Szauter said.
Further development of the technology, in particular the chip design, hinges on additional funding as ElectroSeq is starting to run out of its seed funding.
Szauter, who used to be a training director for the fruit fly database FlyBase at UNM and previously was a bioinformatics professional at the Jackson Laboratory, provided the company with initial funding.
The firm is now hoping to raise on the order of $3 million for the next two years of chip development and has already seen interest from venture capital firms in the Albuquerque area. In parallel, it plans to apply for government funding through the Small Business Innovation Research program.
ElectroSeq is not planning to commercialize its sequencing technology on its own but would seek to be acquired eventually, Szauter said.
Once funded, the next step will be to build a chip with 100 million wells and working fluidics, which would allow the firm to start sequencing DNA. This would be followed by a 1-billion-well chip, a goal the company could reach within two years of being funded, Szauter said, and would enable it to sequence a human genome for $1,000.
Ultimately, ElectroSeq wants to build a 10-billion-well chip. "That's the magic number – that gives us 10 human genomes in a single run at a cost of $100 per genome," Szauter said. He noted that the signal-to-noise ratio might increase and problems with crosstalk might arise at this well density, which he said no one has explored before. "When you go to 10 billion [wells], you're really going into the unknown."
For comparison, the Ion Proton PI chip, which is commercially available, has 165 million wells, and the PII chip, which is still in development, will have 660 million wells. Ion Torrent has repeatedly delayed the commercialization of the PII chip, citing challenges associated with the data transfer rate last year.
In another difference from Ion Torrent's platform, ElectroSeq plans to use reversible terminators instead of unmodified nucleotides, which would help it avoid accuracy problems with homopolymers that both Ion Torrent and 454 ran into. Szauter said the company is currently looking into the cost and intellectual property associated with reversible terminators, a technology Illumina uses as part of its sequencing chemistry. "If we can use reversible terminators … we are going to have the accuracy, the sensitivity, and the throughput," he said.
Szauter said he does not believe ElectroSeq's sequencing chips would violate any patents held by Ion Torrent on pH-based DNA sequence detection, adding that the company's approach will be "sufficiently different" from Ion Torrent's.
Read length would probably be on the order of 200 bases, putting the platform into the short-read technology camp, but preliminary experiments in Edwards' lab have shown that paired-end reads from rolonies should be possible.
While it is waiting for funding to come through, the company is collaborating with another local startup, Bioprocess Diagnostics, on microfluidics for its chips. "We are making dummy chips in the different sizes, so we will be able to do experiments to design, test, and ultimately manufacture the fluidics caps for the 100-million- and the 1-billion-well chips," Szauter said.