Tissue specific cDNAs will be rapidly mapped to mouse chromosomal segments in order to facilitate the definition of both mouse and human genomic organization. An interspecific mouse cross between Mus spretus and a laboratory inbred strain will allow definition of cDNA RFLV markers and mapping by haplotype analysis of most if not all cDNA clones. Equalized neonatal thymus specific cDNA libraries will be obtained by the application of a novel solid phase system in which cDNAs will be bound and eluted from immobilized genomic sequences. This technique should allow the generation of a library of tissue specific cDNAs that cross hybridize between mammalian species by the selection mouse cDNAs that bind to human genomic sequences. Unique cDNAs (approximately 1000) will then be mapped after PCR amplification of sequences. A panel of 38 interspecific backcross mouse DNAs that has been characterized for over 45O markers throughout the mouse genome will be used for rapid mapping. This panel divides the mouse genome into over 470 different segments. A stratified mapping approach will be subsequently utilized to divide the genome into 1000 different segments. In addition, reference markers at 5 cM intervals that have been analyzed in 100 to 400 meiotic events will further define the mapped location of new cDNA markers. The results will provide a putative human regional localization by the application of comparative mapping knowledge. Selected cDNAs, a total of 200, will be used to precisely define the borders of conserved linkage groups by in situ hybridization on human chromosomes. The cDNA clones will also be partially sequenced to provide sequenced tagged sites, allow PCR screening of YAC libraries, and enable the mapping of single stranded conformational polymorphisms by interested investigators. Since 5' as well as 3' sequence information will be obtained, these transcripts will be compared with previously characterized genes. Novel transcripts will provide a resource for molecular genetic studies of thymocyte maturation. In addition, the mapping of over 1000 unique neonatal thymus transcripts may determine whether there are regions of the genome that are transcriptionally active in a coordinate fashion. The mapped cDNA clones will be made easily available to other investigators and should dramatically facilitate closure of long range restriction maps as well as provide additional tools for contig construction of gene rich regions of both the mouse and human genome. The cDNA map will also be integrated with the developing microsatellite STS maps in other laboratories. This will allow integration of efforts to define a contig of the mouse genome.
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