Faculty Research 1990 - 1999

PCR-amplification of simple sequence repeat variants from pooled DNA samples for rapidly mapping new mutations of the mouse.

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Chromosome-Mapping, Crosses-Genetic, DNA: ge, Female, Genes-Recessive, Genetic-Markers, Male, Mice: ge, Mice-Mutant-Strains: ge, Models-Genetic, Mutation, Polymerase-Chain-Reaction: mt, Repetitive-Sequences-Nucleic-Acid, SUPPORT-U-S-GOVT-P-H-S, Variation-(Genetics)

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Genomics 1994 Jun;21(3):626-32


GM18684/GM/NIGMS, CA33093/CA/NCI


A novel strategy for rapidly mapping new mutations of the mouse is presented and tested. The inbred Mus musculus castaneus strain CAST/Ei is used as a tester stock. Separate DNA pools are made from mutant and wildtype intercross (F2) progeny and analyzed using simple sequence repeat (SSR) variants detected using PCR amplification. Linkage is indicated if the mutant DNA pool is enriched for the mutant-strain SSR allelic fragment in comparison with the wildtype pool and controls. The expected contributions of the tester stock to mutant and wildtype pools as a function of recombination frequency are derived for both intercross and backcross progeny segregating for a recessive mutant gene. The sensitivity and reliability of this method is investigated using arbitrary DNA mixtures and also mutant and wildtype pools from a (CAST/Ei x MEV) intercross segregating for the dilute coat color mutation. It is concluded that linkages as great as 20 cM can be reliably detected by this method provided that sufficient progeny contribute to the pools. A set of 39 SSR variants, resolvable by agarose gel electrophoresis and suitable for pooled-SSR analysis, is identified. These are expected to screen 94% of the autosomal genome in crosses between CAST/Ei and common laboratory strains.

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