International Mammalian Genome Society


The 13th International Mouse Genome Conference
October 31-November 3, 1999

Table of Contents * Structure * Bioinformatics * Sequence * Mapping * New Tools * Gene Discovery * Developmental * Mutagenesis * Functional Genomics

D3 Generating a Single Nucleotide Polymorphism (SNP)-Based Genotyping Mapping Panel for Closely Related Strains of Inbred Mice

Al Charest, Barbara M. Jordan, Justin P. Blumenstiel, Julie A. Park, Viktoria Kheifets, John E. Landers and David Housman. Center for Cancer Research, Massachusetts Institute of Technology, 77 Massachusetts Ave., Cambridge, MA, 02139

The current technologies for genotyping inbred strains of mice rely heavily on costly and time consuming manipulations. We have devised a novel approach that relies on principles that greatly alleviate these constraints. First, the complexity of the genome is substantially reduced by creating a number of short DNA fragments that represent the entire genome. This is accomplished in a single PCR reaction with a single oligonucleotide using Interspersed Repetitive Sequence (IRS)-PCR and Degenerate Oligonucleotide Primed (DOP)-PCR strategies. Second, we have found that homologous IRS- and DOP-PCR fragments from different inbred strains of mice contain sufficient base substitutions or SNPs to be used as a means of discrimination between different strains. Genotyping of each SNP is carried out using Allele Specific Oligonucleotide (ASO) hybridization using a standard condition for all probes. The generation and characterization of a panel of >150 SNP-based markers covering the entire genome will be discussed. These markers were typed for 96 different strains of inbred mice thus creating the potential to select a set of markers that will discriminate alleles between any pair of strains. The applications of SNPs-based genetic markers in genetic linkage and Loss of Heterozygosity studies will also be discussed. SNPs are an abundant class of genomic polymorphisms which hold great promise as a tool for genotyping. Coupling genome complexity reduction strategies with SNP-derived ASO hybridization technology permits the determination of large numbers of genotypes in a single experiment using standard lab equipment. We have developed a SNP- based genotyping technology which circumvents the need for large numbers of PCR reactions and repetitive optimization procedures. This highly efficient and low cost genotyping system will invariably facilitate the mapping and isolation of modifier loci and quantitative trait loci between closely related strains of mice.

 


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