The long-term objective is a nanopore detector chip for a general utility instrument capable of inexpensive de novo sequencing that can also be used for re-sequencing projects. The instrument directly generates base-dependent electronic signals as multi-kilobase length fragments of single stranded genomic DNA is driven sequentially through nanopores articulated with electrically contacted single walled carbon nanotube probes. The final system is intended to provide a relatively high quality sequence from =6.5-fold coverage of a genome using DNA from fewer than 1 million cells, with no amplification or labeling.
The specific aims are: 1) Characterize ungapped nanotube articulated nanopore detectors in ionic solution with and without DNA molecules to establish a device model; 2) Control ssDNA binding, translocation, and sliding on the nanotube surface exposed in ungapped nanotube articulated nanopores; 3) Study and optimize DNA molecule induced field effect modulation of nanotube electrode conductance in ungapped nanotube articulated nanopores as a function of nanotube bias, gate voltage, and solution properties; 4) Analyze and optimize tunneling current modulations between gapped nanotube electrodes in the first generation detector; 5) Design and fabricate a second generation detector with embedded 'T' nanotube geometry and achieve 1 Kb/sec sequencing on Kb length strands of DNA; 6) Design a third generation nanopore detector for high throughput 10 Kb/sec/nanopore sequencing. If we are able to resolve each base as it passes through a nanopore at the rate of 104 bases/sec as proposed here, an instrument with an array of 100 such nanopores could produce a high quality draft sequence of one mammalian genome in ~20 hours at a cost of approximately $1,000/mammalian genome. Genomic sequencing at these reduced costs would make vital contributions to improved human health on many fronts, including the understanding, diagnosis, treatment, and prevention of disease; environmental science and remediation; and the genetics of human health and disease derived from the understanding of evolution. PROJECT
We are developing the core detector of an instrument that could produce a high-quality draft sequence of one mammalian genome in ~20 hours at a cost of approximately $1,000/mammalian genome. Genomic sequencing at these reduced costs would make vital contributions to improved human health on many fronts, including the understanding, diagnosis, treatment, and prevention of disease; environmental science and remediation; and the genetics of human health and disease derived from the understanding of evolution. ? ? ?
| Fleming, Stephen J; Lu, Bo; Golovchenko, Jene A (2017) Charge, Diffusion, and Current Fluctuations of Single-Stranded DNA Trapped in an MspA Nanopore. Biophys J 112:368-375 |
| Deamer, David; Akeson, Mark; Branton, Daniel (2016) Three decades of nanopore sequencing. Nat Biotechnol 34:518-24 |
| Levine, Edlyn V; Burns, Michael M; Golovchenko, Jene A (2016) Nanoscale dynamics of Joule heating and bubble nucleation in a solid-state nanopore. Phys Rev E 93:013124 |
| Rollings, Ryan C; Kuan, Aaron T; Golovchenko, Jene A (2016) Ion selectivity of graphene nanopores. Nat Commun 7:11408 |
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| Szalay, Tamas; Golovchenko, Jene A (2015) De novo sequencing and variant calling with nanopores using PoreSeq. Nat Biotechnol 33:1087-91 |
| Hoogerheide, David P; Lu, Bo; Golovchenko, Jene A (2014) Pressure-voltage trap for DNA near a solid-state nanopore. ACS Nano 8:7384-91 |
| Nagashima, Gaku; Levine, Edlyn V; Hoogerheide, David P et al. (2014) Superheating and homogeneous single bubble nucleation in a solid-state nanopore. Phys Rev Lett 113:024506 |
| Hoogerheide, David P; Albertorio, Fernando; Golovchenko, Jene A (2013) Escape of DNA from a weakly biased thin nanopore: experimental evidence for a universal diffusive behavior. Phys Rev Lett 111:248301 |
| Lu, Bo; Hoogerheide, David P; Zhao, Qing et al. (2013) Pressure-controlled motion of single polymers through solid-state nanopores. Nano Lett 13:3048-52 |
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