This application is directed at """"""""Pilot Projects or Feasibility Studies for Genomic Analysis"""""""" (PA-90-21) in support of the Human Genome Initiative (HGI), and seeks to develop two-dimensional (2D) gels of DNA fragments as a method to detect and resolve up to several thousand independent genomic segments in a single gel. Such an ability would enable at least an order- of-magnitude improvement in the rapidity of producing saturated genetic and physical maps of any organism, and would greatly facilitate multiple types of genomic analysis through the ability to simultaneously examine the segregation or other status of large numbers of markers distributed across an entire genome. The proposed study will employ the mouse as its experimental mammalian model due to the wealth of available inbred and well-characterized genetic stocks; however, the methods developed should be generally applicable with little further modification to any genetic system. In brief, existing techniques of 2D electrophoresis of DNA fragments (by size and base composition/sequence), probing for highly repetitive DNA sequences, computer-assisted analysis of complex spot patterns and classical mouse genetics will be combined in a novel synthesis. The repetitive sequences to be explored will include both previously characterized repeats such as Intracisternal A Particles (IAPs), which are widely distributed throughout the mouse genome and are highly polymorphic both within and between inbred strains, and oligonucleotides representing subsets of LINE-1 elements. A single blot probed with one of these sequences should enable simultaneous analysis of hundreds or even thousands of independent genomic segments. Such a blot could be reprobed as desired to reveal the distribution of additional families of repetitive sequences, and individual spots could be recovered for analysis of unique flanking regions and determination of a generally accessible Sequence Tagged Site (STS) from each resolved copy of a repetitive sequence.

Agency
National Institute of Health (NIH)
Institute
National Human Genome Research Institute (NHGRI)
Type
Exploratory/Developmental Grants (R21)
Project #
5R21HG000845-02
Application #
2209084
Study Section
Genome Study Section (GNM)
Project Start
1992-06-08
Project End
1995-05-31
Budget Start
1993-06-01
Budget End
1995-05-31
Support Year
2
Fiscal Year
1993
Total Cost
Indirect Cost
Name
University of Cincinnati
Department
Genetics
Type
Schools of Medicine
DUNS #
City
Cincinnati
State
OH
Country
United States
Zip Code
45221