Our major goal is to understand how genes control the structure, function, and development of the nervous system in a vertebrate. Initially, we plan to give the highest priority to the isolation and study of mutations which affect the structures of photoreceptor cells in the retina of the eye of the zebra fish (Brachydanio rerio.) A corollary goal of our work is to develop rapid and efficient procedures that will permit a variety of genetic analyses in a vertebrate at the level of the whole organism. Heterozygosity in diploid eukaryotes often makes genetic studies cumbersome. We have perfected methods for making homozygotes in zebra fish: Sperm whose genetic contribution has been eliminated by ultraviolet irradiation are used to activate eggs. The maternal haploid set of chromosomes is allowed to replicate once, and partitioning of the duplicated chromosomes into two cells is prevented by hydrostatic pressure or heat shock. A cell with two identical sets of chromosomes is thereby produced. We propose to mutagenize mother fish during their early development with ethylnitrosourea (ENU). Among the progeny, clones of mutants will be recognized by use of a vision-dependent feeding test. Mutants will be characterized through optomotor and optokinetic tests, flicker fusion frequency, electroretinogram shape, and retinal anatomy. In addition to the direct screening of mutagenized progeny, gamma-ray-induced mutations (possibly long deletions) with specific retinal lesions will be used to select ENU-induced point mutations. Through the study of these mutants we hope to learn what kinds of alterations are specific to retinal photoreceptor cells, what the developmental, structural, functional, and behavioral consequences of the mutations are, how mutation affects other cell types, and what the molecular nature of the mutations are.
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