Hybridization-Assisted Nanopore DNA Sequencing

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DESCRIPTION (provided by applicant): The objective of the proposed research program is to develop a low-cost nanopore sequencing technology taking advantage of the latest advances in solid-state nanopore technology and utilizing the recently proposed concept of hybridization-assisted nanopore sequencing (HANS). It has been proposed by the PI and coworkers that by hybridizing short matching strands of oligonucleotides, known as DNA probes, and using nanopore ionic conductance to detect the positions of the probes, one can construct DNA sequences using the ionic current profiles, thus avoiding the classical "repeat problem" in the standard sequencing-by-hybridization (SBH) method, and bypassing the harsh requirement of single-base spatial resolution (0.4nm) in the original nanopore sequencing proposal of Kasianowicz et al. The proposed HANS sequencing technology has the potential of being fast, low-cost and portable. The proposed research is to test the feasibility of the HANS concept. The specific aims of the project are: (1) Determine the spatial resolution of the ionic conductance method using a combined optical tweezers and nanopore setup. (2) Parallel manipulation of DNA translocations in multiple nanopores. (3) Detect hybridization of single DNA probes on a single-stranded ssDNA using optical tweezers and ionic conductance of the nanopore. (4) Re-sequencing a sample DNA sequence to determine accuracy. (5) Develop non-specific DNA-bead binding technology for de novo sequencing. The proposed research will greatly advance the field of bio-nanotechnology and could directly result in a low-cost DNA sequencing technology, which in turn will have far-reaching benefits to the public health of the US in disease detection and in the studies of the molecular processes underlying diseases. The research program provides excellent training opportunities for students and postdoctoral researchers in preparing them to join the workforce of the emerging bio-nanotechnology economy.