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

D4 Large-Scale Detection and Genotyping of Mouse Single-Nucleotide Polymorphisms

K. Lindblad1, N. Patil2, E. Winchester1, D. Wang1, E. Robinson1, M. J. Daly1, N. Shah2, J. Warrington1, T. J. Hudson1,3, E. Lander1. 1Whitehead Institute/MIT Center for Genome Research, Cambridge, MA; 2Affymetrix Inc, Santa Clara, CA; and 3Dept of Medicine and Human Genetics, Montreal General Hospital Research Institute, McGill University, Canada

A single nucleotide polymorphism (SNP) is a position in the genome where two alternate bases each occur at an appreciable frequency in a population. SNPs are abundant in mammalian genomes and amenable to automated genotyping. SNPs are especially well suited to genotyping of mouse crosses: While such bi-allelic markers are less informative than microsatellites in humans, they are completely informative in a cross of inbred mouse strains. In order to generate a set of useful SNPs for the mouse genome, we performed a screen of 3717 mouse sequence tagged sites (STSs) in a DNA panel of 8 inbred strains using high-density variation detection GeneChip probe arrays. Altogether, 2848 SNPs were found in 1755 STSs (325 ESTs and 1428 random sequences). Not surprisingly, the majority of the SNPs detected were variations between Mus musculus castaneus (CAST/Ei) and the seven lab strains screened. This resulted in a mean SNP frequency of 1/202 bp between M. m. castaneus and lab strains. Among lab strains a mean pairwise SNP frequency of 1/1029 bp was found. Sixty-eight percent of the STSs containing SNPs had previously been assigned map positions. A map was constructed yielding a mean inter-marker distance of 2.6 cM as an average for M. m. castaneus versus lab strains. The pair-wise mean inter-marker distance among lab strains was 7.5 cM. To test the usefulness of these SNPs, 100 well-spaced SNPs between A/J and C57BL/6J were chosen and a genotyping panel developed. The SNPs were assayed by PCR amplification in two pools of 50 loci, followed by genotyping by single-base extension (SBE) in 6 pools of 16-17 loci and detection in 6 lanes on an ABI 377. By loading multiple times on a gel, a cross of 48 mice can be genotyped for 100 markers on two gels. This effort is a first step towards a high-density SNP map of the mouse, as well as an efficient system for genotyping SNPs, that will be useful in positional cloning of single gene traits as well as for dissection of complex traits.

 


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