KOMP2-Phase 2 Production and Phenotyping by the DTCC Consortium

KOMP 2-DTCC联盟的第2阶段生产和表型分析 摘要 本申请是为了竞争性地延长DTCC财团参与第二阶段的资金， 敲除小鼠生产和表型分析（KOMP 2）项目。DTCC联盟成员中心 包括加州戴维斯大学（UCD;牵头机构），表型基因组学中心（TCP）， 多伦多、儿童医院奥克兰研究所（CHORI）和查尔斯河实验室 International Inc（CRL）.我们建议利用我们的经验，并继续使用经过良好考验的协调 成功实施KOMP 2-第1阶段的管理、战略、方案和流程， 扩大和加强KOMP 2-第2阶段的活动。在KOMP 2-第1阶段，DTCC已完成（或正在进行） 在第1阶段结束时完成所有既定目标并实现所有指标（例如，加速显微注射 胚胎干细胞克隆，生殖系突变体生产率提高，生产时间轴提前完成， 跟踪完成表型分析），超出了我们工作范围所需的目标和目的， 由DTCC在KOMP 2-Phase 2中提出。在这次更新中，DTCC建议将男性和女性的表型 使用以下方法产生的至少1，500个突变纯合（HOM）或杂合（HET）小鼠系的队列： Cas9 RNA引导的核酸酶（Cas9 RGN）和相应的年龄和性别匹配的野生型（WT）对照， 3个表型分析管道（胚胎、幼年和成年），使用所有IMPC要求的和几个可选的IMPReSS- 在多个器官系统中建立测试。在应用基于中心的过程质量保证之后， 产品（小鼠和数据）的质量控制，所有数据（包括元数据）将立即上传到 数据协调中心（DCC）和小鼠表型信息基础设施（MPI 2）， 公众可通过IMPC门户网站（www.impc.org）查阅。提高生成鼠标的效率 线路和数据，DTCC成员中心将进行技术开发项目，以改善和增加 生产效率（例如高效RGN介导的靶向，lacZ敲入插入）和先导 表型分析试验（例如，自动化饲养笼监测和微核[基因组不稳定性]），以确定 在我们的青少年和成人管道中的附加值，可重复性和成本效益。表型 技术开发活动将包括一种新的定量试剂盒为基础的蛋白质组学测定，和代谢组学 分析突变小鼠品系，有望增加重大的科学价值，并提高研究的实用性， 表型数据。此外，DTCC将通过寻求提名积极参与科学界的工作 对于要研究的基因（例如，Adult Pipeline），在生产时优先响应KOMP 2小鼠的请求， 表型分析正在进行中，并在存入突变小鼠资源和研究中心后进行 （www.mmrrc.org），并提供增值服务和协作（例如，PAR-13-231表型分析 胚胎致死，Baylor-Harwell KOMP 2-Phase 2联盟的组织病理学支持）。除了输入 从NIH项目官员，DTCC将指导DTCC小组的意见和建议， 专家顾问、NIH KOMP 2科学顾问委员会和IMPC科学顾问小组。的 DTCC将参加所有KOMP 2和IMPC联盟活动，包括每月电话会议， 小组委员会工作组和会议、国家和国际会议、实地考察和其他 相关活动。为了最大限度地提高科学产出和对生物医学研究的影响，DTCC还 承诺通过以下方式与这些生产、表型分析和研发工作协调， 其他KOMP 2资助的参与者。总而言之，到5年结束时，DTCC将为 基因型确认的小鼠和至少1，500个突变小鼠品系的经验证的广泛表型数据 及时并在预算范围内，以支持KOMP 2计划和IMPC在功能上注释 哺乳动物基因组

KOMP2-Phase2 Production and Phenotyping by the DTCC Consortium ABSTRACT This application is to competitively renew funding for the DTCC Consortium’s Phase2 participation in the Knockout Mouse Production and Phenotyping (KOMP2) Project. Member centers of the DTCC Consortium include the University of California Davis (UCD; lead institution), The Centre for Phenogenomics (TCP) in Toronto, the Children’s Hospital Oakland Research Institute (CHORI), and Charles River Laboratories International Inc (CRL). We propose to leverage our experience and continue to use well-tested coordinated management, strategies, protocols, and processes successfully implemented for KOMP2-Phase1 to increase, expand, and enhance activities for KOMP2-Phase2. In KOMP2-Phase1, the DTCC completed (or is on track to complete by the end of Phase1) all established goals and achieve all metrics (e.g., accelerated microinjection of ES cell clones, increased production rate of germline mutants, early completion of production timeline, on- track for completion of phenotyping) that exceed objectives and goals required for the scope of work we propose by the DTCC in KOMP2-Phase2. In this renewal, the DTCC proposes to phenotype male and female cohorts for at least 1,500 mutant homozygous (HOM) or heterozygous (HET) mouse lines produced using Cas9 RNA-guided nuclease (Cas9 RGN) and corresponding age and sex –matched wildtype (WT) controls in 3 phenotyping pipelines (Embryo, Juvenile, and Adult) using all IMPC-required and several optional IMPReSS- established tests across multiple organ systems. After applying center-based quality assurance of processes, and quality control of products (mice and data), all data (including meta-data) will be uploaded without delay to the Data Coordination Center (DCC) and then the Mouse Phenotyping Informatics Infrastructure (MPI2) for public access through the IMPC web portal (www.impc.org). To enhance the efficiency of generating mouse lines and data, DTCC member Centers will conduct technology development projects to improve and increase efficiency of production (e.g. high-efficiency RGN-mediated targeting, lacZ knockin insertion) and pilot phenotyping tests (e.g. automated home cage monitoring and micronucleus [genome instability]) to establish their added-value, reproducibility, and cost-effectiveness in our Juvenile and Adult Pipelines. Phenotyping technology development activities will include a novel quantitative kit-based proteomic assay, and metabolomic profiling of mutant mouse lines that promise to add significant scientific value and increase research utility of phenotyping data. In addition, the DTCC will actively engage the scientific community by seeking nominations for genes to study (e.g., Adult Pipeline), prioritize responses to requests for KOMP2 mice while production and phenotyping is in progress and after deposition into the Mutant Mouse Resource and Research Center (www.mmrrc.org), and provide value-added services and collaborations (e.g., PAR-13-231 Phenotyping Embryo Lethals, histopathology support to the Baylor-Harwell KOMP2-Phase2 Consortium). In addition to input from NIH Program Officers, the DTCC will be guided by advice and recommendations of the DTCC Panel of Expert Advisors, the NIH KOMP2 Scientific Advisory Board, and the IMPC Panel of Scientific Consultants. The DTCC will participate in all KOMP2 and IMPC consortium activities, including monthly teleconferences, subcommittee working groups and meetings, national and international conferences, site visits, and other relevant activities. In order to maximize the scientific output and effect on biomedical research, the DTCC also commits to coordinate its efforts with those production, phenotyping, and research and development efforts by other KOMP2-funded participants. In summary, by the end of 5 years, the DTCC will have contributed genotype-confirmed mice and validated broad-based phenotyping data for at least 1,500 mutant mouse lines on time and within budget in support of the KOMP2 program and the IMPC effort to functionally annotate the mammalian genome.