CLONING THE POLAR LETHAL OVUM MUTANT GENE OF THE MOUSE

描述(改编自《调查者摘要》)：当女性 DDK近交系与许多其他自交系的雄性交配，最高可达 95%的胚胎在植入前发育过程中死亡。这个 DDK雄性与其他近交系雌性之间的正反交， 是完全可行和肥沃的。额外的遗传分析表明 致死性状分离为两个紧密连锁的基因座，新命名为Omo (DDK综合征的母亲特有成分)和OMS(父亲 Omo起作用的基因)。不同寻常的父母出身和 这一现象的基因型依赖性质(即 母系Omo-DDK因子与非DDK父系等位基因紧密连锁 基因座导致了这些胚胎的死亡，但相互的组合 不受影响)使隔离的因素(S)对此负责 几个领域的工作者感兴趣的行为，包括遗传学， 胚胎学和基因组印迹。私家侦探构建了一个基因 包含11号染色体区域的精细结构和物理图谱 OMO和OMS，并建议使用他们收集的资源来隔离 编码母体因子的基因导致了“DDK综合征”。 他们的位置克隆方法很简单，将包括三个 具体目标：1)鉴定一个或多个酵母人工染色体 (YAC)通过显微注射获得含有OMO基因某些部分的克隆 YAC介导的杂交种耗尽DDK卵RNA进入“野生型”胚胎。DDK卵子 已耗尽Omo(Omo基因-a的RNA产物)的RNA 已在DDK卵子细胞质中发现的致死RNA)有望 对野生型胚胎的活力没有影响。2)完成 细菌人工染色体(BAC)文库的筛选 BAC介导的微量注射分离和鉴定 杂交种耗尽的DDK OVA RNA进入包含以下内容的克隆的“野生型”胚胎中 奥莫基因。3)BAC介导的杂交选择法分离Omo基因 DDK OVA文库的构建或DDK OVA文库的筛选 阳性BAC的单拷贝DNA片段。

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.