High-resolution cosmid mapping of the left arm of Saccharomyces cerevisiae chromosome XII; a first step towards an ordered sequencing approach.

For the sequencing of the left arm of chromosome XII of Saccharomyces cerevisiae, we fine-mapped the entire 450 kb fragment between the ribosomal DNA (rDNA) and the left telomere. Total yeast DNA in agarose blocks was digested with I-PpoI, which exclusively cuts once in each repeat unit of the rDNA. The resulting fragment was isolated from pulsed-field gels, together with the equally sized chromosome IX. A cosmid library of some 30-fold chromosome coverage was generated from this material, with the cloning efficiency being around 20,000 clones per microgram genomic DNA. The chromosome XII and IX specific clones were identified by complementary hybridizations with the respective chromosomes. For the left arm of chromosome XII, a contiguous cosmid array (contig) with an average map resolution better than 9 kb was generated by clone hybridization procedures. The ordered library serves as a tool for the physical mapping of genetic markers. Also, a minimal set of 15 clones was selected that covers the entire fragment. This subset forms the basis for the generation of a template map of much higher resolution for a directed sequencing of the left arm of chromosome XII.