STRUCTURAL INVESTIGATION OF NOVEL MUTANT HEMOGLOBINS

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The studies proposed in this application are to provide a focused and integrated approach, through a genome center, for the construction of high-resolution and comprehensive genetic and physical maps of human chromosome 13. The Pittsburgh Human Genome Research Center (PHGRC) represents the cooperative and collaborative efforts of several investigators at the University of Pittsburgh, Carnegie Mellon University, and the Wistar Institute to produce a genetic linkage map at an average resolution of 1-2 megabase pairs (Mbp) and a physical linkage map at an average resolution of 200-250 kilobase pairs (kb) for human chromosome 13. Our strategy is to construct these maps using sequence tagged sites (STSs) from clones containing polymorphic simple sequence repeats, other anonymous sequences, and DNA sequences. By means of in situ hybridization, DNA probes containing simple sequence repeats from chromosome 13 will be mapped into 11 ``bins" approximately 10 Mbp each. To achieve a uniform genetic linkage map, approximately 500 STSs constructed from these clones, and other clones containing cDNA or anonymous sequences, will be ordered using radiation hybrids. Yeast artificial chromosome (YAC) clones corresponding to the ordered STSs will be identified and assembled into contigs. These resources will be used to construct high-resolution genetic and radiation hybrid maps; ordered YAC contigs covering 96% or more of chromosome 13 will also be obtained. The construction of new tools, novel fluorophors for labelling DNA molecules and new computing tools for rapid and automated mapping, are essential components of our research proposal that will have wide utility for investigators focusing on other chromosomes and organisms. The outcome of these studies will be a set of molecular, genet ic and computational tools, and a high-resolution map of human chromosome 13. Ultimately these studies will lead to the elucidation of the corresponding homologous regions in the mouse. Our studies will provide resources that can lead to the rapid recovery of disease genes localized to chromosome 13.