Discovery of Craniofacial and Retinal Genes

描述(由申请人提供)：针对RFA DE-03-001“颅面部疾病的基因发现”，本申请是为了在斑马鱼中发现一个独特的高效基因发现计划，以寻找人类疾病基因的候选基因：我们的具体目标是发现斑马鱼基因，当斑马鱼基因发生突变时，会导致(1)颅面部骨骼发育不全，(2)视网膜退化。对于两者来说，都将利用一种强大的基因发现方法的新组合。基因中的损伤将通过病毒介导的插入突变引入。由于病毒插入用于标记突变的基因，因此可以极快地识别和克隆这些基因，通常在几天内就能完成。突变体将在高效、定向的第一代(G1)筛选中进行鉴定，使用通过孤雌生殖获得的胚胎和幼虫，并用荧光染料进行重要染色来揭示表型。孤雌生殖和G1筛选消除了数月的时间，并节省了饲养数十万条鱼所需的时间，这是更常见的正向遗传策略，即提高突变体家系和筛选F3后代。新的品系将迅速提供给其他使用社区资源的研究人员，斑马鱼信息网络将为突变体做广告，斑马鱼国际资源中心将分发它们。斑马鱼胚胎和幼虫的各种有利属性将使详细的表型研究和快速表征基因在面部骨骼和眼睛发育中的功能成为可能。

DESCRIPTION (provided by applicant): In response to RFA DE-03-001 "Gene Discovery for Craniofacial Disorders" this application is for a unique program of efficient gene discovery in zebrafish to find candidates for human disease genes: Our specific aims are to discover zebrafish genes that when mutated result in (1) craniofacial bone dysgenesis, and (2) retinal degeneration. For both, a novel combination of powerful approaches toward gene discovery will be utilized. Lesions in genes will be introduced by viral-mediated insertional mutagenesis. Because the viral insertions serve to tag the mutated genes, the genes can be identified and cloned extremely rapidly, often within a few days. The mutants will be identified in highly efficient, directed, first generation (G1) screens, using embryos and larvae obtained by parthenogenesis and vitally stained with fluorescent dyes to reveal the phenotypes. Parthenogenesis and G1 screening eliminates months of time and saves rearing hundreds of thousands of fish required in the more usual forward-genetic strategies of raising families of mutants and screening F3 progeny. The new strains will be made rapidly available to other researchers using community resources, the Zebrafish Information Network to advertise the mutants and the Zebrafish International Resource Center to distribute them. Various favorable attributes of the zebrafish embryo and larva will permit detailed phenotypic studies and rapid characterization of the functions of the genes in facial bone and eye development.