We propose generating a complete molecular physical, cytogenetic and genetic linkage map of human chromosome 13 (CH-13). The physical map will be based on positioning YAC and cosmid contigs on CH-13, including cytogenetic localization of the contigs. The ends of the CH-13 YAC's and cosmids will be sequenced to provide STS's at spacings of 0.1 Mb or less. The map will be enriched by assigning macro-restriction sites (e.g. Not I) and anonymous cDNA's to the physical and cytogenetic maps. The technologies needed to accomplish these goals have been established in our preliminary studies. The CEPH YAC library is being used as the source for CH-13 specific YAC's, which we are identifying by screening with PCR probes derived from radiation hybrids of hamster: human monochromosomal 13 cell lines. Regional cytogenetic assignments of CH-13 YAC's and cosmids are being made by assaying with deletion panels and in situ hybridization. The YAC and cosmid contigs are being generated by screening of dense filters of the arrayed libraries and a cross- hybridization algorithm, as well as by end-clone walking. CH-13-specific cDNA's are being identified by screening of normalized human liver and brain libraries and a novel sub-cloning strategy. cDNA assignments are being made by cytogenetic mapping, as well as by localization to specific members of YAC and cosmid contigs. The genetic linkage map is based on polymorphic CA repeats and tri- and tetra-nucleotide repeats, at a resolution of less than 2 cM. The genetic linkage map is being constructed by typing reference pedigrees and statistical analysis. All of the map data will be incorporated into discrete and relational databases to super-position all of the physical, cytogenetic, and genetic findings into an integrated map. The evolving reagents and data will be made available via an outreach program to all qualified investigators both national and international. We anticipate completing the CH-13 map within the five years of requested support.
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| Fischer, S G; Cayanis, E; Russo, J J et al. (1994) Assembly of ordered contigs of cosmids selected with YACs of human chromosome 13. Genomics 21:525-37 |
