Genome function in Mouse with Combination Mutagenesis

描述（由申请人提供）：该提案寻求3年的支持，以完成小鼠5号染色体50 Mb区域的功能分析，该区域占小鼠基因组的近2%。在以前的资助下，通过用乙基亚硝基脲（ENU）随机诱变基因组来诱导突变，并使用利用称为rump-white（Rw）的染色体倒位的育种方案选择近端Chr 5上的突变。创建了一组跨越大部分Rw区域的缺失复合物以帮助这些突变的遗传分析。尽管对几种表型进行了筛选，但胚胎致死突变（总共34个，代表32个互补组）是回收的最大类别。此外，恢复了两个导致不育的男性突变，一个导致耳聋，两个影响行为。致命的突变作用于整个发育过程，从植入前到出生。突变体表型包括：内细胞团增殖失败;异常胎盘形成;原肠胚形成缺陷;类稳态骨骼转化;颅面异常;和心血管缺陷。遗传作图实验已经将这些突变中的大多数定位到Rw区域内相对较小的间隔。迄今为止，已克隆了5个致死和1个不育等位基因。这项建议有两个具体目标。第一个是定位克隆剩余的不育突变和20个致命等位基因。第二个是使用适合于不同胚胎学阶段的分析方法完成该致死突变集的总表型表征。结合起来，这些实验将分子缺陷与突变表型联系起来，从而阐明正常哺乳动物发育的遗传机制。许多未克隆的突变目前映射到不包含或很少包含已知致死基因的区域，确保几个新的必需基因或功能元件将被鉴定。表型/基因型关系的建立将为我们和哺乳动物发育界进行更详细的研究铺平道路。总的来说，这些实验最终将提供一个公正的一瞥到基因组的代表性区域编码的功能元件。该项目使用小鼠作为模型，以确定影响人类胚胎发育和生育能力的重要基因。最终，我们预计这项工作将有助于治疗某些男性不育症，增加我们对正常胚胎发育所需基因的理解，并可能改善某些出生缺陷的基因诊断。

DESCRIPTION (provided by applicant): This proposal seeks 3 years of support to complete functional analyses of a 50 Mb region of mouse Chromosome 5 comprising nearly 2% of the mouse genome. Under previous funding, mutations were induced by randomly mutagenizing the genome with ethylnitrosourea (ENU), and selecting those on proximal Chr 5 using a breeding scheme that exploited a chromosome inversion called rump-white (Rw). A set of deletion complexes spanning much of the Rw region was created to aid in the genetic analysis of these mutations. Though screens were conducted for several phenotypes, embryonic lethal mutations (total of 34, representing 32 complementation groups) were the largest class recovered. Additionally, two male mutations causing infertility, one causing deafness, and 2 affecting behavior were recovered. The lethal mutations act throughout development, from pre-implantation through birth. The mutant phenotypes include: failure of the inner cell mass to proliferate; abnormal placental formation; gastrulation defects; homeotic-like skeletal transformations; craniofacial abnormalities; and cardiovascular defects. Genetic mapping experiments have localized most of these mutations to relatively small intervals within the Rw region. Thus far, 5 lethals and 1 infertility allele have been cloned. This proposal has two specific aims. The first is to positionally clone the remaining infertility mutation and 20 of the lethal alleles. The second is to complete a gross phenotypic characterization of this lethal mutation set, using analytic methods appropriate for various embryological stages. In combination, these experiments will link the molecular defects to mutant phenotypes, and thereby illuminate genetic mechanisms of normal mammalian development. Many of the uncloned mutations currently map to regions containing none or few known lethal genes, ensuring that several new essential genes or functional elements will be identified. This establishment of phenotype/genotype relationships will pave the way for more detailed investigations by ourselves and the mammalian development community. Overall, these experiments will eventually provide an unbiased glimpse into the functional elements encoded by a representative region of the genome. The project uses the mouse as model to identify important genes affecting embryonic development and fertility in humans. Ultimately, we anticipate this work will assist treatment of certain male infertilities, increase our understanding of the genes required for normal embryonic development, and potentially lead to improved genetic diagnosis of certain birth defects.