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Japan's Quantum Biosystems Shows Raw Read Data from Single-Molecule Nanogap Sequencer

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Quantum Biosytems of Japan, which has been developing a single-molecule electronic DNA sequencing technology that measures tunneling currents from DNA bases as the molecule passes by a sub-nanometer gap, publicly showed single-read raw sequence data for the first time this week at the Personalized Medicine World Conference in Mountain View, Calif.

Quantum is now in the midst of raising additional funding as well as awareness of its technology and plans to start an early-access program for a prototype sequencer this year.

"We are going to increase our global exposure in order to hire talented researchers and engineers, and also to get some funding from outside of Japan," Toshihiko Honkura, the company's president and CEO, told In Sequence.

Based in Osaka, Quantum Biosystems was founded in early 2013 by Honkura and Masateru Taniguchi, a professor at the Institute of Scientific and Industrial Research at Osaka University and the firm's chief technology officer. The company has 12 full-time employees and a number of external advisors, including Tomoji Kawai, a professor at Osaka University, and Yoshinobu Baba, a professor at Nagoya University.

Quantum has exclusively licensed intellectual property from both universities related to statistical methods for analyzing tunneling currents to make base calls, controlling the speed of DNA translocating nanopores, and extending single-stranded DNA using so-called nanopillars, and it closely collaborates with researchers at Osaka University. In addition, the firm has developed its own patented or patent-pending technologies for manufacturing nano-silicon devices as well as for noise reduction.

So far, Quantum has raised nearly $5 million in total, including $1.3 million in seed financing from Japanese venture capital firms and $3.5 million in grants from the Japanese government. It is in the midst of another venture financing round, with $5 million already secured from Japanese VC firms and plans to raise at least another $5 million from VCs outside of Japan.

Additional funding would allow Quantum to accelerate its development. "We'd like to put a lot of money behind scaling out the platform," said Nava Whiteford, the company's executive officer for informatics, who joined Quantum last year from Oxford Nanopore Technologies, where he was a principal computational scientist.

Whiteford said he was impressed by "how far they have developed the technology with relatively little cash and in a relatively short space of time."

Quantum's core technology relies on a pair of nanoelectrodes that are separated by a sub-nanometer gap, across which it places a bias voltage. It then detects changes in the tunneling current as single-stranded DNA or RNA translocates through the gap.

The company currently fabricates the nanoelectrodes by placing a nanowire on a piece of silicon and forming a gap. Using this so-called mechanically controlled break junction, "we can get reasonably decent reads," Whiteford said.

The next generation of the technology, which the company is developing in parallel, uses a gating nanopore to confine the motion of translocating DNA, as well as prefabricated nanogaps.

Last year, researchers led by Osaka University's Taniguchi and Kawai published a paper in Scientific Reports in which they showed that they can distinguish the four types of DNA and RNA bases by their tunneling currents and that they can resequence trinucleotides and reconstruct the sequence of slightly larger oligonucleotides.

Although those data showed the DNA bases can be measured with single-base precision, the DNA moved randomly around the nanoelectrodes and "you could not actually generate useful reads from that technology," Whiteford said.

In the meantime, the researchers have come up with a way to control the motion of DNA by electrophoresis, using "the same basic mechanism as gel electrophoresis," he said.

This has allowed them to generate reads of about 20 base pairs from short oligonucleotides with an accuracy of more than 99 percent in non-homopolymer regions, and around 90 percent in regions with homopolymers.

"Considering the amount of time and effort that has been put into this technology, I feel like this is a huge achievement," Whiteford said. "We're very confident in the quality of our data."

Whiteford explained that Quantum decided to show its data early in order to give scientists a chance to evaluate it, unlike Oxford Nanopore Technologies, for example, which has chosen not to show any raw sequence data publicly yet.

"We want to see how this initial data release is received publicly," Whiteford said. "Our hope is that we'll get some engagement from the community, and the fact that we're trying to make an effort to be open will be well received. If everything goes well, we'll release more data in the coming weeks."

The data quality is "good enough" to sequence the 5.4-kilobase genome of bacteriophase phi X 174, he said, a commonly used test genome, and there is room to improve the DNA motion control and to reduce the noise.

One of the main differences between protein nanopore-based systems like Oxford's and Quantum's technology is that the latter measures individual bases, whereas protein nanopores obtain a signal from several bases at once and need to deduce individual bases from that. "The challenge [for protein nanopores] has always been to somehow increase that resolution so you get single-base resolution," Whiteford said.

Stuart Lindsay, a professor at Arizona State University whose team has also worked on DNA sequencing by measuring tunneling currents, told In Sequence that the controlled translocation of DNA has been a "big issue" for his group, so "it will be interesting to find out how this new company gets around that problem."

He said he is not sure how an intact single-stranded DNA molecule, which is about 1 nanometer wide, can pass through a sub-nanometer gap.

Whiteford told In Sequence that the company adjusts a gap size of initially 0.5 nanometers during the experiment until it sees a good quality signal, which often happens at an estimated gap size of about 0.8 nanometers. In that case, the DNA likely passes "very near the electrodes, but not through the smallest gap," and it still generates a tunneling current.

Besides improving its technology further, Quantum will need to scale up its devices, which are currently built at Osaka University. Using the current technology, "we can build a device integrating more than 10s of channels, and there is no technical limitation to integrating thousands to millions of channels," Whiteford said.

According to Honkura, Quantum already has a preliminary partnership with an unnamed Japanese high tech company to manufacture silicon devices.

"Getting the R&D production process that we use to develop the devices at the moment and pushing that into a commercial facility is one of the things that we have to hit," Whiteford said, and "more cash would help alleviate that problem because it helps speed things up."

In addition, he said, the firm wants to forge partnerships with researchers and companies outside of Japan. For example, Quantum plans to start an early-access program for a prototype of its sequencing technology this year, which is "by invitation only," according to its website. Commercialization could start as early as next year, Whiteford said.

While he hesitated to estimate the specifications of a commercial sequencer, the ultimate goal is to build an instrument that can generate "hundreds of gigabases per hour" with "potentially very long reads," he said. The data quality will not drop off along the reads, so "it's all down to controlling the motion of the DNA."

"We're not really aiming at a niche market," Whiteford said. "We definitely think we can compete both on volume and quality," as well as on the ability to detect methylation and to sequence RNA directly.

The price of the commercial sequencer has not been determined yet, but it will likely cost less than $10,000 to build, Honkura said, noting that the current prototype costs about $30,000 to produce. The disposable silicon devices currently cost on the order of $50, which could go down to less than $1 with mass production, he said.

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