ISOLATION AND CHARACTERIZATION OF BOVINE CHROMOSOME REGION-SPECIFIC SEQUENCE-TAGGED MICROSATELLITE SITES (STMS)

A strategy to fill in gaps in small targeted regions of bovine chromos omes is presented. The procedure involved microdissection of subchromo some areas, their PCR amplification and cloning. Out of three approach es which have been tested to amplify chromosome material two have show n to be not suitable for generation of bovine chromosome region specif ic probes. In contrast, probes generated by DOP-PCR after treatment wi th Topoisomerase I showed region specific rehybridization on the corre sponding dissected chromosome areas indicating a representative and co mplex pattern of specific sub chromosomal sequences. Nevertheless, all methodical steps still require special emphasis on accurate handling to exclude formation of artifacts and contamination with DNA of other sources. Using this approach chromosome region specific libraries were established from the chromosome areas 1q1.1-1.4, 5q2.1-2.4, 1q1.3-2.4 and 12q2.4-2.6 providing an average of 0,06-0,6% microsatellite conta ining several of which were of human. Three of the isolated bovine spe cific microsatellites revealed polymorphism (FBN5 and FBN8 from chr 5q 2.1-2.4; FBN7 from chr 1q1.3-2.4); one of them (FBN5) was genotyped in the IBRP and assigned to its predefined subchromosomal origin by link age analysis.