By Julia Karow
This article, originally published Feb. 27, has been updated from an earlier version to include additional information and comments.
MARCO ISLAND, Fla. — Ion Torrent Systems unveiled an electronic sequencer last week that reads DNA on a semiconductor chip by measuring the release of hydrogen ions as nucleotides get incorporated by DNA polymerase.
The instrument will cost less than $50,000 and generate "hundreds of millions of bases" and "millions" of highly accurate reads per run, each several hundred bases in length, according to Jonathan Rothberg, the company's co-founder and CEO. Each run will take about an hour and cost less than $500.
Speaking in front of a packed audience at the end of the last session of the Advances in Genome Biology and Technology conference here, Rothberg said that the company, which is based in Guilford, Conn., and San Francisco and has been operating quietly since its foundation in 2007, plans to sell tens of thousands of the instruments to laboratories around the world.
Although the system uses polymerase-based sequencing-by-synthesis chemistry — like most existing second-generation sequencers — it is the first to do away with lasers, cameras, or labels, relying entirely on electronic detection. "The machine is now a chip," Rothberg said.
This allows the company to piggyback on decades of investment and development by the semiconductor industry, and to produce its proprietary chips in standard CMOS factories. "Our sequencers are built in the same factories that make the parts for your iPhone, your camera, and your computer," he said. "Four hundred foundries in the world can make our wafers, and there are 1,500 packages that serve the electronic industry that can package our chips."
At the heart of the desktop-size sequencer is a 9 by 9 millimeter semiconductor chip, currently consisting of an array of about 1.55 million 3.5-micrometer wells, an ion-sensitive layer underneath, and one electronic sensor per well below.
The company has already developed a future generation of the chip, shrinking the wells to 1.3 micrometers, increasing the number of wells and sensors to several hundred million, and speeding up DNA synthesis to less than two seconds per base, from the current rate of four seconds per base.
To sequence DNA, amplified template is loaded into the wells, along with DNA polymerase. Reminiscent of 454's sequencing chemistry, unmodified nucleotides are then sequentially added to the system and incorporated by the enzyme at a rate of four seconds per base. Each base incorporation releases hydrogen ions, which decreases the pH of the surrounding solution. "Our sequencer is, essentially, the world's smallest solid-state pH meter," Rothberg explained.
The change in pH charges the ion-sensing layer. The charge is then converted into a voltage, which is read by the sensors at a rate of 60 times per second, so more than 200 data points are collected for each base in four seconds. This high-speed sampling, coupled with a high signal-to-noise ratio, leads to "extremely high accuracies," Rothberg said, and will allow for reads of "well over" 500 bases.
So far, the platform has provided perfect reads of 100 to 200 bases, Rothberg said, adding that with software improvements, there is "no reason" why it should not be able to produce highly accurate 500-base reads in the future.
In homopolymer stretches, the system generates a signal that is proportional to the number of repeated bases, similar to the 454 platform, which initially had high error rates in these regions. However, Rothberg said that Ion Torrent's system, because the electronic signal remains linear over a wide range, can accurately detect stretches of up to six identical bases and possibly even longer ones. "We think it will go as far as biologically relevant," he said.
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Rothberg provided no details about the sample-preparation method but said that the system could use "every library you have in your refrigerator," including libraries prepared for the Illumina, SOLiD, and 454 platforms.
In principle, Ion Torrent could use, for example, a bead-based emulsion-PCR amplification method, like SOLiD and 454, or a "nanoball" amplification approach like Complete Genomics.
Sequencing reagents — DNA polymerase and natural nucleotides — cost on the order of a dollar, though packaged kits will be more expensive than that, he said.
The instrument, which Ion Torrent workers carried on stage as Rothberg gave his talk, was built with diagnostic applications in mind, and is manufactured at an ISO 13485-certified facility, meaning "it's ready to be FDA-registered at your clinic," he said. "You guys find amazing content, so we can immediately put these machines in any clinic down the hall and use your content to change peoples' lives."
Rothberg provided no timeline for a commercial release of the system, although the company said last week that it plans to launch its "Personal Genome Machine" sequencer this year.
In a proof-of-concept experiment in collaboration with the Broad Institute, Ion Torrent sequenced the 34-kilobase adenovirus genome at 59-fold coverage, dubbed by the Broad's Chad Nusbaum the "first post-light genome." The technology covered 100 percent of the genome with a consensus accuracy of 99.9999 percent, equivalent to a quality score of 61.
The company has also sequenced the 4.6-megabase E. coli genome at 12.9-fold coverage, covering more than 99 percent of the genome with more than 99.9 percent accuracy. Rothberg said that the sequence showed no systematic error and that coverage across the genome was even.
In addition, Ion Torrent has sequenced 841 megabases from a large genome — 3,123 exons from 207 genes — and has other collaborative work in progress. Rothberg listed both the Broad Institute and the Stanford Genome Technology Center as collaborators.
On the data analysis side, Ion Torrent is partnering with four bioinformatics firms at the moment — CLC Bio, DNAStar, Geospiza, and Partek — who are still working on optimizing algorithms for Ion Torrent's data. DNAStar generated the assembly for the E. coli genome.
Ion Torrent is currently looking for additional scientific collaborators and plans to give away two of its instruments under a grant program to researchers who suggest “killer applications” for the system (see other article, this issue).
Following Rothberg's talk, audience members uniformly said they were intrigued by the new technology, though many cautioned there were unanswered questions, especially regarding the sample prep, which Rothberg said he would address in a future talk.
George Grills, director of operations of core facilities at Cornell University's life sciences core laboratories center, said a $500 sequence run that can be completed in one hour would be "pretty amazing" and would create a "sweet spot" for the system. "It's like having a 454 on your desktop," he said.
According to Mike Snyder, professor of genetics at Stanford University School of Medicine, Ion Torrent's "principle is promising." He said he liked the fast turnaround time and the relatively long reads, compared to other platforms.
But scientists are waiting for more data and details about the technology to emerge. "The proof is in the pudding," Snyder said.