HIGH-RESOLUTION GENE MAPPING BY IN SITU HYBRIDIZATION

点击翻译按钮获取中文摘要

The overall objective of this project is to facilitate the construction of detailed physical and genetic maps of human and murine chromosomes. We will continue ongoing work with the CEPH/Genethon group to complete a low resolution YAC contig map of each human chromosome by using FISH techniques to analyze the 800 YAC contigs (approx. 85% genome coverage) assembled to date to verify chromosomal location, to determine chimeric clone content and to identify false contigs. Orphan YACS (with no contig assignment and no STS marker) also will be FISH mapped as will several hundred contigs identified with genetically defined STSs so that a direct correlation between the genetic and cytogenetic maps can be made. When, and if necessary, individual YAC clones from contigs exhibiting chimerism and false joints will be FISH mapped separately to identify and weed out problem clones. In collaboration with MIT Genome Center, we will jointly construct a low resolution YAC contig map of the mouse genome. We will also prepare, arrange and distribute chromosome-specific YAC sublibraries that will be made by screening the genome YAC library with high complexity PCR products generated from normal and Robertsonian fusion mouse chromosomes. High resolution maps will be constructed for murine chromosome 1, regions of human and mouse chromosomes containing zinc finger gene clusters, as well as other chromosomal loci containing known disease genes or interesting gene families. We will provide a FISH mapping service to any investigator in the genome community who has a newly identified, but unmapped gene, or a novel transgenic mouse where information on chromosomal locus of the transgene would be informative. Finally, we will fully automate our digital imaging microscopes, both to increase the throughput of FISH experiments by a factor of 10 fold and to permit the multiplex screening of high density, gridded arrays of YAC clones with combinatorially labeled fluorescence probes.