The Jackson Laboratory Knockout Mouse Production and Phenotyping Project (JAX KOMP2)

项目摘要/摘要 老鼠和人类有大约20,000个基因。到目前为止，其中一半以上的数据还很少 基因和近三分之一没有功能注释。因为它们之间的高度相似 在小鼠和人类的基因集合中，小鼠产生的遗传数据往往可以推断为人类的基因 功能。小鼠和人之间具有共同功能的基因的小鼠模型可以导致新的 人类疾病，这对药物筛选、临床前研究和加深对生物学的理解是有用的 和致病机制。Knokout鼠标表型程序(KOMP2)的目标是生成线条 携带全基因组小鼠基因敲除(KO)的小鼠，并使小鼠 基础广泛的表型。杰克逊实验室(JAX)提议将其目前资助的KOMP2 小鼠的生产和冷冻保存(U42)和敲除小鼠表型(U54)操作作为 RFA-RM-15-017有限竞赛：淘汰小鼠生产和表型项目(UM1)。 JAX KOMP2建议使用尖端且经济高效的Cas9 RNA引导核酸酶(Cas9 RGN，又称Cas9 RGN 称为CRISPR/Cas9)技术，用于在整个过程中产生、繁殖、冷冻保存和表型为1500株的小鼠 项目期。将继续努力改进Cas9 RGN技术，以降低成本， 提高靶向效率，创造更复杂的突变等位基因。基因将在协调中进行选择 与我们的KOMP2和IMPC合作伙伴，并将重点放在那些注释不佳的基因上，这些基因具有显著的 社区需求和与其他NIH支持计划的整合，以及预测在选择中起作用的基因 小路。为了保证随时进入社区，我们将把老鼠送到外面的调查人员那里 被活生生地放在货架上，并将线条存入老鼠突变区域资源中心(MMRRC) 供将来使用的储存库。 对18周龄以下的幼年动物进行广泛的表型鉴定将在所有1500个品系上进行 使用国际小鼠表型联盟(IMPC)所需和JAX特定协议的小鼠。我们会 评估体重和成分、代谢和生理参数以及行为和认知 功能。为了检测年龄相关的表型，我们将使用相同的管道来检测20%(总共300)的表型 15-18个月龄之间的线条。根据KOMP2当前阶段产生的数据，我们 预计大约30%的线路将不可行。我们将使用高通量来表征不能存活的突变体 三个胚胎时间点的成像方式。所有由胚胎、幼年和成年小鼠产生的数据 将迅速存入支持KOMP2和IMPC的数据协调中心(DCC)。 最后，日本宇宙航空研究开发机构将与KOMP2区域网络和 IMPC分享协议、创新和新技术，并广泛和公开地传播我们的发现 通过出版物、在会议上的介绍、网络活动和社交媒体向国际社会宣传。

PROJECT SUMMARY/ABSTRACT Mice and humans share approximately 20,000 genes. To date, little data exists for more than half of these genes and nearly one third have no functional annotation. Because of the high degree of similarity between the mouse and human gene set, genetic data generated in mice can often be extrapolated to human gene function. Mouse models of genes with common functionality between mice and men can lead to new models of human disease, which are useful for drug screening, preclinical studies and deeper understanding of biological and disease mechanism. The goal of the Knockout Mouse Phenotyping Program (KOMP2) is to generate lines of mice that carry knockouts (KOs) for a genome-wide collection of mouse genes and subject the mice to broad based phenotyping. The Jackson Laboratory (JAX) proposes to merge its currently funded KOMP2 Mouse Production and Cryopreservation (U42) and Knockout Mouse Phenotyping (U54) operations as part of RFA-RM-15-017 Limited Competition: Knockout Mouse Production and Phenotyping Project (UM1). JAX KOMP2 proposes to use cutting-edge and cost-effective Cas9 RNA-guided nuclease (Cas9 RGN, also called CRISPR/Cas9) technology to generate, breed, cryopreserve and phenotype1500 lines of mice during the project period. Continued effort will be made to improve the Cas9 RGN technology so as to reduce costs, increase targeting efficiency, and create more complex mutant alleles. Genes will be selected in coordination with our KOMP2 and IMPC partners, and will focus on those with poor annotation, genes that have significant community demand and integrate with other NIH-support programs, and genes predicted to function in select pathways. To guarantee ready access to the community, we will ship mice to outside investigators while they are alive on the shelf and deposit the lines into the Mouse Mutant Regional Resource Center (MMRRC) repositories for future use. Broad based phenotyping on juvenile animals up to 18 weeks of age will be performed on all 1500 lines of mice using International Mouse Phenotyping Consortium (IMPC)-required and JAX-specific protocols. We will assess body weight and composition, metabolic and physiological parameters, and behavioral and cognitive function. To detect age-dependent phenotypes, we will use the same pipeline to phenotype 20% (300 total) of the lines between 15-18 months of age. Based on data generated from the current phase of KOMP2, we expect about 30% of lines to be non-viable. We will characterize the non-viable mutants using high-throughput imaging modalities at three embryonic time points. All data generated from embryonic, juvenile and adult mice will be rapidly deposited into the Data Coordination Center (DCC) that supports KOMP2 and the IMPC. Lastly, JAX will work collaboratively with the KOMP2 Regional Network and with member organizations of the IMPC to share protocols, innovation, and new technology and to broadly and openly disseminate our findings to the international community through publication, presentations at meetings, web activities and social media.