CLONING THE POLAR LETHAL OVUM MUTANT GENE OF THE MOUSE

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DESCRIPTION (Adapted from the Investigator's Abstract): When females of the DDK inbred strain are mated with males of many other inbred strains, up to 95% of the resulting embryos die during preimplantation development. The reciprocal crosses, between DDK males and females of other inbred strains, are fully viable and fertile. Additional genetic analysis indicates that the lethal trait segregates as two closely-linked loci, newly designated Omo (the maternal-specific component of the DDK syndrome) and Oms (the paternal gene upon which Omo acts). The unusual parental-origin and genotype-dependent nature of this phenomenon (i.e. the combination of a maternal Omo-DDK factor with a non-DDK paternal allele at the closely-linked locus results in the death of these embryos, but the reciprocal combinations are unaffected) makes the isolation of the factor(s) responsible for this behavior of interest to workers in several fields, including genetics, embryology and genome imprinting. The PI has constructed a genetic fine-structure and physical map of the region of chromosome 11 that contains Omo and Oms and proposes to use the resources they have assembled to isolate the gene encoding the maternal factor responsible for the "DDK syndrome". Their positional cloning approach is straightforward and will include three specific aims: 1) Identification of one or more yeast artificial chromosome (YACs) clones that contain some portion of the Omo gene by microinjection of YAC-mediated hybrid-depleted DDK ova RNA into "wild-type" embryos. DDK ova RNA that has been depleted for Omo (the RNA product of the Omo gene - a lethal RNA that has been identified in DDK ova-cytoplasm) is expected to have no effect on the viability of wild-type embryos. 2) Completion of the screening of a bacterial artificial chromosome (BAC) library and the isolation and characterization (by microinjection of BAC-mediated hybrid-depleted DDK ova RNA into "wild-type" embryos) of clones that contain the Omo gene. 3) Isolation of an Omo cDNA by BAC-mediated hybrid selection of a DDK ova cDNA library or screening of a DDK ova cDNA library with single-copy DNA fragments of a positive BAC.