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Home > Press > New Nanopore Technique Facilitates Faster, Cheaper Genome Analyses

Schematic of a solid state nanopore used for genome analyses (not to scale). The electrostatic potential near an approximately five nanometer-wide, solid-state nanopore attracts negatively-charged, double-stranded DNA molecules into the pore, which electronically detects the molecules as they traverse the pore. (Photo courtesy of Nature Nanotechnology.)
Schematic of a solid state nanopore used for genome analyses (not to scale). The electrostatic potential near an approximately five nanometer-wide, solid-state nanopore attracts negatively-charged, double-stranded DNA molecules into the pore, which electronically detects the molecules as they traverse the pore. (Photo courtesy of Nature Nanotechnology.)

Abstract:
Ultra-fast, low-cost genomic sequencing and profiling may some day accelerate the pace of biological discovery and enable clinicians to quickly and precisely diagnose patients' susceptibility to disease and tolerance of selected drugs. But this scenario may not be realized until engineers find a way to considerably increase the sensitivity of sensors used to detect the DNA molecules that define the human genome.

New Nanopore Technique Facilitates Faster, Cheaper Genome Analyses

Boston, MA | Posted on December 21st, 2009

It's a feat that could be achieved by reducing the number of target DNA molecule copies needed to obtain an accurate read. And that presents a formidable challenge: to produce sufficient copies to decipher the genome using current technology, most scientists still rely on time-consuming, expensive, and error-prone DNA replication tools such as the polymerase chain reaction (PCR).

Now researchers have devised a method that advances the prospects for efficiently analyzing DNA samples without amplification. In a study published in the Dec. 20 online edition of Nature Nanotechnology, Associate Professor Amit Meller (BME, Physics), BME postdoctoral fellow Meni Wanunu, BU physics student Will Morrison and collaborators at New York University and Bar-Ilan University demonstrated a method to tune solid-state nanopores tiny, nearly cylindrical, silicon nitride sensors that electronically detect DNA molecules as they pass through the pore to require far fewer DNA molecules than ever before.

"This study shows that using our method, we can detect a much smaller amount of DNA than previously published," said Meller. "When people will start to implement genome sequencing or profiling using nanopores, they could use our nanopore capture approach to greatly reduce the number of copies used in those measurements."

Nanopore capture consists of two distinct steps: the arrival of a sample molecule to the pore mouth, and the threading of the end of that molecule into the pore. To significantly increase the rate at which nanopores capture incoming, two nanometer-wide DNA molecules, Meller and his colleagues used salt gradients to alter the electric field in the pore's vicinity. This achieved a funneling effect that directed charged DNA molecules toward the mouth of the pore and boosted the molecules' arrival and threading rates.

By upping the capture rate by a few orders of magnitude and decreasing the volume of the sample receiving chamber, the researchers reduced the number of DNA molecule copies required for nanopore-based detection by a factor of 10,000 from about one billion sample molecules to 100,000. They also demonstrated that longer DNA molecules (containing tens of thousands of nucleotide base pairs) increased the capture rate even further.

"PCR and other DNA replication technologies limit DNA molecule length," said Meller. "Because our method avoids amplification, it not only reduces the cost, time and error rate of DNA replication techniques, but also enables the analysis of very long strands of DNA."

Funded by the National Institutes of Health and the National Science Foundation, the research team set out to achieve a better understanding of the physical forces that govern the DNA capture process. They arrived at their findings by using high-end transmission electron microscopes (TEM) to fabricate hundreds of nanopores with atomic-scale precision, and testing differently configured salt gradients near the pores.

"We had to perform extensive studies with these nearly atomic-scale pores in order to reveal how the electrostatic potential, which extends at least hundreds of nanometers away from the pore, focuses DNA into and through the pore," said Meller.

To conduct further investigations of unamplified genomes, Meller is now exploring other technologies, including optical detection and force measurements, for reading single DNA molecules as they pass through nanopores.

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