Citation: N/A Interpretive Summary: Microbes containing large fragments of mammalian DNA have proven to be very useful tools for gene mappers, however, they also have limitations. This manuscript describes the difficulties that must be dealt with when using these "large- insert genomic libraries" for gene mapping. Nine bovine genetic markers have been genetically mapped because of their association with specific genes on a large-insert clone. Previous bovine mapping data and comparative mapping information suggested that all of these new microsatellites would map to bovine chromosome 15. The 9 new markers mapped to a total of 7 different chromosomes, none of which was chromosome 15. Possible explanations are discussed.
Technical Abstract: Two strategies were used to increase the microsatellite marker density and contribute to the comparative map of a meat tenderness QTL region on bovine chromosome 15. In the first strategy, PCR primer pairs were designed to amplify fragments of genes predicted to map to BTA15 (APOA1, CCKBR). PCR products were sequenced and analyzed using BLAST to confirm specific amplification. The PCR products were radioactively labeled and used to probe a BAC library. Positive BACs were identified and verified by secondary hybridization and or PCR. BAC DNA was extracted, subcloned and screened for microsatellites. The second strategy utilized markers previously shown to map to BTA15 to screen a YAC library by PCR. YACs were isolated that contain each microsatellite and subcloned into Bluescript. Subclones were screened for new microsatellites. Primers sets were designed using subclone sequences to amplify microsatellites in genomic DNA. The segregation of alleles for all markers was consistent with autosomal Mendelian inheritance except for BY1501 which maps to the non-psuedoautosomal region of the X chromosome. The original expectation was that all markers derived from these clones would map to BTA15, based on comparative analysis with human chromosome 11 or previous bovine mapping data. Indeed, a number of markers isolated in the overall study did map to BTA15 and are described elsewhere. The remaining markers are described here and are useful as markers on their respective chromosomes.