Sequence it ... and they will come

May 18, 2012

Rapid DNA sequencing may soon become a routine part of each individual’s medical record, providing enormous information previously sequestered in the human genome’s 3 billion nucleotide bases. This week’s NEWSFOCUS section of the journal Science describes recent advances in sequencing technology.

Stuart Lindsay, director of the Biodesign Institute’s Center for Single Molecule Biophysics, is on the forefront of this research, having successfully addressed a central stumbling block in nanopore sequencing – reading single nucleotide bases in a DNA chain. Lindsay’s latest experimental results, which demonstrate critical improvements in DNA reads,  have just appeared in the journal Nanotechnology. Download Full Image

Once accurate sequencing falls below the threshold of $1,000 per genome, the technology should become ubiquitous, according to many. As the current Science overview suggests, that day may be drawing near as both the speed and cost of whole genome sequencing advances at a pace outstripping Moore’s famous Law, (which dictates a doubling of computing power – and halving of the expense – every 18 months).

The latest technological competition involves the idea of threading a single strand of DNA through a tiny, molecular-scale eyelet known as a nanopore. This strategy may soon allow the entire DNA sequence to be read in one go, rather than cut apart, deciphered in brief fragments and painstakingly re-assembled.

While the first sequencing of the human genome took researchers 13 years and $3 billion to achieve, under the auspicies of the Human Genome Project, the feat may soon be accomplished at the blinding  rate of 6 billion nucleotide bases every 6 hours at a cost of $900. At least that is the extravagant claim being made by Oxford Nanopore Technologies, one of the pioneering companies driving new sequencing developments.

Since the seemingly quixotic idea of nanopore sequencing was first thought up in the mid 1990s, enormous advances have been made. The basic idea is that when a nanopore is immersed in a conducting fluid and a voltage is applied across it, conduction of ions through the nanopore will produce a measurable electric current. This current is highly sensitive to the size and shape of the nanopore and in theory, each nucleotide base in the DNA thread will obstruct the nanopore as it migrates, altering the ionic current in a recognizable and reproducible way.

The DNA “thread” is tricky material to manipulate however – so fine that it would take about 5000 DNA strands laid side by side to equal the width of a human hair. Just finding a suitable eyelet at this scale proved a challenge. At first, porous, transmembrane proteins were explored. Alpha hemolysin (αHL), a bacterium that causes lysis of red blood cells, seemed a particularly promising candidate, given the nanopore diameter required for sequencing DNA.

Since then, other protein-based portals for DNA have been tinkered with and more recently, various “solid state” nanopores of silicon or graphene have been investigated. These can be more easily fabricated and their properties, more precisely controlled.

According to Science’s review of the present state of the art, nanopore sequencing “seems poised to leave the lab,” and the dream of a $1000 genome may be close at hand, though challenges remain. A persistent problem in sequencing individual bases has been that they tend to stream through the nanopore too rapidly to pinpoint each base independently. Instead, the measured current in early experiments reflected the average produced by a group of bases wending their way through the tunnel.

Lindsay’s technique relies on reading electrical current in a tiny circuit composed of a DNA nucleotide trapped between a pair of gold electrodes, which span a nanopore. The electrodes are made by functionalizing the tip of a scanning tunneling microscope (STM), with molecules that can bind individual DNA bases as they poke their heads through the nanopore.

Recognition Tunneling, the name Lindsay applies to his sequencing method, relies on outfitting one of two electrodes with sensing chemicals, the other with the nucleotide target to be sensed. A signal is produced when the junction between sensing chemical and target self-assembles, closing the circuit.

In this type of junction, where lengths separating electrodes are down to a molecular scale, electrons can exhibit odd behavior associated with the quantum subatomic world, “tunneling” through barriers under conditions prohibited by classical physics.  In such a scenario, each of the 4 nucleotides should produce a signature tunneling current, which can be used to sequence DNA base-by-base as it feeds its way through the nanopore. Trapping each base momentarily allows time for an accurate identification, before it is released and the DNA thread continues its transmigration through the nanopore.

Replacing ionic current flow with tunneling current can potentially improve sequencing resolution considerably and in their latest work, Lindsay’s group demonstrates that multiparameter analysis of the current spikes produced by tunneling can indeed identify each DNA base as it is temporarily pinned by hydrogen bonding between the functionalized electrodes.

There’s more.

In addition to pinpointing nucleotide identity with greater than 90 percent accuracy, the technique also permits environmental gene modifications to be identified, for example, methylation. This represents a major advance for sequencing, as such epigenetic alterations to the genome have profound implications for the study of human health and disease, including embryonic and post-natal development, and cancer.

The Nanotechnology paper describes a new approach to analyzing the tunneling signals.  The Lindsay group used machine learning (the process used by IBM’s Watson to win at Jeopardy) to train a computer to recognize the DNA bases.  The machine called all four bases (A,T,C and G) as well as the “fifth base” – methyl – that carries the epigenetic code, with 96 percent accurarcy on a single molecule read.

“Oxford Nanopore have a made a tremendous breakthrough in nanopore sequencing using ion current, as highlighted in the NEWSFOCUS story,” Lindsay says.  “But we think we can bring even more to the table with the supersensitivity and chemical resolution of Recognition Tunneling.”

Roche Pharmaceuticals has recently licensed the technology.

The high stakes race for rapid sequencing appears to be entering the home stretch, though new surprises are likely before the finish line. Once it is crossed, the era of personalized medicine will have arrived. Many new insights into the genomic basis of human health and disease are almost certain to follow.

Stuart Lindsay is also a professor in the Department of Chemistry and Biochemistry in the College of Liberal Arts and Sciences

Richard Harth

Science writer, Biodesign Institute at ASU


US Energy Secretary visits ASU for research briefing

May 18, 2012

Energy research at Arizona State University was the focus of a visit by U.S. Energy Secretary Steven Chu on May 15. Chu, who was in the Valley of the Sun to help launch the second phase of the Solar Phoenix program, stopped by the university’s Tempe campus to meet with ASU President Michael M. Crow, LightWorks director Gary Dirks and a number of scientists engaged in DOE funded research.

Chu is charged with helping implement President Obama's ambitious agenda to invest in clean energy, reduce our dependence on foreign oil, address the global climate crisis, and create millions of new jobs.  Download Full Image

Chu, a distinguished scientist and co-winner of the Nobel Prize for Physics in 1997, has devoted his recent scientific career to the search for new solutions to energy challenges. Previously, Chu was the director of the Department of Energy's Lawrence Berkeley National Lab, where he led the lab in pursuit of alternative and renewable energy technologies. Chu, a member of the National Academy of Sciences as well as several other academies around the world, has a doctorate in physics from the University of California, Berkeley.

While at ASU he talked briefly about the mission of the Department of Energy, highlighting ARPA-E, energy and science as “touch points.” But Chu’s main purpose in coming to ASU was to listen.

In describing ASU as a New American University, Crow explained that “we have set ourselves apart from other major public universities by having a differentiated set of design aspirations. For instance, use-inspired scholarship, valuing entrepreneurship, merging disciplines, and fueling things in a different kind of way. It’s a different approach.”

Also speaking at the gathering that was held in ASU’s newest Interdisciplinary Science and Technology Building (ISTB4) was Dirks, the former president of BP Asia-Pacific and BP China. Dirks is director of LightWorks, an ASU initiative that capitalizes on the university’s strengths in solar energy and other light-inspired research, including the Arizona Center for Algae Technology and Innovation. 

“We try to cover energy in an integrated way and the Department of Energy is an important supporter of ours,” said Dirks, who also is a distinguished sustainability scientist with the Global Institute of Sustainability, a strategic research unit of ASU’s Office of Knowledge Enterprise Development.

Among the scientists who briefed Chu on DOE-sponsored projects were Christiana Honsberg, a professor in the School of Electrical, Computer and Energy Engineering, part of ASU’s Ira A. Fulton Schools of Engineering, and Cody Friesen, an associate professor in the School for Engineering of Matter, Transport and Energy.

Honsberg leads a national Engineering Research Center at ASU that is supported jointly by the National Science Foundation and Department of Energy to solve challenges to harnessing solar power in economically viable and sustainable ways. Known as QESST – Quantum Energy and Sustainable Solar Technologies – the center focuses on identifying alternative energy sources that solar power can provide through photovoltaic technologies.

“The general approach of QESST is that photovoltaics can meet the terawatt challenge within the next decade or two, provided we have advances that sustain the growth. The central driving force is that there is absolutely fundamental new physics across the entire span of the different technologies,” said Honsberg.

Friesen leads a research team pursuing advances in battery technology and energy storage. Specifically, the research group is developing a new type of ultra-high-energy metal-air batteries that use advanced ionic liquids, and promise to provide low-cost, long-range power for all-electric and hybrid vehicles. In the long run, this advance could significantly reduce the need for the United States to import oil since more of the energy to power transportation could be drawn from the nation’s electrical grid.

The work being done by Friesen and his research team is an example of how ASU is “leveraging the government’s money and accelerating the movement of this technology,” according to Crow. 

Among the other ASU faculty members attending the briefing included Willem Vermaas, a professor in ASU’s School of Life Sciences and the Center for Bioenergy and Photosynthesis. Vermaas’ research team is working on a form of photosynthetic bacteria, called cyanobacteria, which are modified to over-produce and secrete fatty acids for biofuel feed stocks using just sunlight, water and carbon dioxide as inputs.

Also in attendance was Devens Gust, a Regents’ Professor in chemistry and biochemistry who oversees an Energy Frontier Research Center. The ASU center, one of 46 EFRCs, is pursuing advanced scientific research on solar energy conversion based on the principles of photosynthesis, the process by which plants convert sunlight to energy.

“Sustainability and renewable energy, advanced materials, flexible systems, and an arrangement of other areas we are concentrated in. We have strengths in research overall,” said Crow.

ASU media contact:
Sharon Keeler

Amelia Huggins

DOE media contact:
Jen Stutsman
202-586-3261 office | 240-364-4727 cell
U.S. Department of Energy, Office of Public Affairs

Britt Lewis

Communications Specialist, ASU Library