INTEGRATED ZEBRAFISH GENOMIC RESOURCES

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DESCRIPTION: (Applicant's abstract) The zebrafish Danio rerio is an important vertebrate model system for mutational analysis of developmental processes. Efficient utilization of the zebrafish requires a variety of genetic and genomic resources not yet fully developed, including expressed sequence tags (ESTs), high density genetic and physical, and robust inbred mapping strains. We propose an integrated approach to genomic resource development, including EST sequencing, that will be used to develop sequence tagged sites (STSs), that will in turn be used to identify single nucleotide polymorphisms (SNPs) between inbred strains, and mapped on radiation hybrid (RH) and meiotic mapping panels. (1) Generation of Expressed Sequence Tags (ESTs) from various stages of zebrafish development. We will generate 5' and 3' sequences from representative cDNAs from each of 50,000 different clusters identified by oligonucleotide hybridization fingerprint analysis of 278,000 independent zebrafish cDNA clones. (2) STS development and Radiation Hybrid mapping. We will generate 10,000 STS markers from 3' EST reads and genotype 5,000 STSs on a Radiation Hybrid panel to construct a marker dense physical map, including STSs with SNPs to allow integration of physical and genetic maps. (3) Single nucleotide polymorphism marker development and Meiotic mapping. We will re-sequence 10,000 EST-based STS from inbred strains used in meiotic mapping to identify Single Nucleotide Polymorphisms. We will type 1800 SNPs at restriction enzyme sites on meiotic mapping panels for generation of a marker dense genetic map. Identification of up to 50,000 zebrafish homologues of human genes by EST sequencing will facilitate functional analyzes of these genes not possible in humans. Moreover, genetic and physical mapping of 10,000 zebrafish EST will provide candidate genes for zebrafish mutations and a dense framework for initiating positional cloning projects. Finally, comparison of map positions of zebrafish and human genomes. This will allow zebrafish mutations to reveal roles for human genes and increase our understanding of the basis of human disease.