The zebrafish is an important model organism for studying vertebrate development, cell biology and cancer. Large-scale mapping and sequencing efforts currently rely on piecing together randomly characterized DNA fragments. Molecular cytogenetic methodologies adds a unique and complementary mapping approach by visualizing the presence of DNA sequences at specific chromosomal sites. Unfortunately, molecular cytogenetic probes for the zebrafish are lacking. Molecular cytogenetic studies will bridge the gap between the physical and genetic maps and expedite the genetic dissection of zebrafish mutant models of human diseases. This application proposes to develop a comprehensive set of molecular cytogenetic tools for the zebrafish by integrating cytogenetic landmarks into the zebrafish genome project. This will be accomplished first by standardizing the zebrafish karyotype and correlating the physical chromosomes with the genetic linkage maps using specific near-centromeric BAC probes. Five hundred BACs will be mapped by fluorescence in situ hybridization (FISH) to wild type metaphase chromosomes and anchored to existing maps and contigs. Information obtained will be posted on an updatable and interactive zebrafish cytogenetic website. The second objective will be to develop chromosome-specific FISH probes and an array CGH platform. BAC probes will be identified which map near the centromere and telomeres of each zebrafish chromosome. These probes will be used to characterize gamma-irradiated mutant lines thought to contain deletions or chromosomal translocations. Also, """"""""paint probes"""""""" for each zebrafish chromosome will be produced and used to characterize interspecific chromosomal rearrangements and supernumerary marker chromosomes. Finally, a 3 Mb resolution BAC-based array CGH platform will be constructed. This tool will be used to identify genomic DNA gains and losses which could lead to the identification of novel oncogenes and tumor-suppresor genes that lead to tumorigenesis in many zebrafish mutants.
