Nanopore detection of haplotype

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DESCRIPTION (provided by applicant): I will develop the technology and basic science needed to use nanopores for high speed SNP detection and haplotyping. The technology depends on driving single polynucleotide molecules through a nanopore coupled to sensitive single channel recording electronics that can provide a direct read-out of the polymer's characteristics. The development of these new molecular diagnostic methods takes advantage of three recent discoveries: (1) A membrane channel, or nanopore, can be used as a high-throughput device that detects and probes single molecules as they translocate through the nanopore; (2) The ionic current through the nanopore is sensitive to local changes in the cross-sectional area of the translocating molecule; (3) A new, planar fabrication method - ion beam sculpting - that allows us to create a single digit nanoscale pore of a desired dimension in robust solid state insulating membranes. Our work will optimize: (a) zinc-finger protein labeling of DNA; (b) the electrical readout of DNA length; (c) the minimum fragment length and number of different zinc-finger proteins needed to achieve reliable SNP identification and high-speed haplotyping. The proposed technologies will impact basic research areas such as development and cancer that must deal with complex sets of genes and mutations, and where existing methods to rapidly examine the linkages between a large number of polymorphic sites on multiple chromosomes in a large number of individuals are limiting. The tools and basic research proposed will open new possibilities for the future development of high sensitivity, information rich diagnostic methods that are critical for early disease detection.