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.

Agency
National Institute of Health (NIH)
Institute
National Human Genome Research Institute (NHGRI)
Type
Research Project (R01)
Project #
5R01HG002338-03
Application #
6765838
Study Section
Genome Study Section (GNM)
Program Officer
Schloss, Jeffery
Project Start
2002-09-30
Project End
2006-06-30
Budget Start
2004-07-01
Budget End
2006-06-30
Support Year
3
Fiscal Year
2004
Total Cost
$507,606
Indirect Cost
Name
Harvard University
Department
Microbiology/Immun/Virology
Type
Schools of Arts and Sciences
DUNS #
082359691
City
Cambridge
State
MA
Country
United States
Zip Code
02138
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King, Gavin M; Schurmann, Gregor; Branton, Daniel et al. (2005) Nanometer patterning with ice. Nano Lett 5:1157-60
Wang, Hui; Dunning, James E; Huang, Albert P-H et al. (2004) DNA heterogeneity and phosphorylation unveiled by single-molecule electrophoresis. Proc Natl Acad Sci U S A 101:13472-7