Transgenic and Targeted Mutagenesis Laboratory Shared Resource

摘要-转基因和靶向诱变实验室 转基因和靶向突变实验室（TTML）是西北大学范围内的资源 致力于基因工程小鼠的产生、进口、回收/再衍生和冷冻保存。 TTML为Lurie癌症中心（LCC）成员提供广泛的服务，包括生成 通过显微注射的转基因、基因编辑和靶向转基因小鼠模型; 胚胎干（ES）细胞;将ES细胞显微注射到胚泡中以产生生殖系感受态嵌合体; 从种质中引进和恢复/再衍生小鼠品系;以及小鼠精子的冷冻保存 和胚胎。自上次审查以来，核心已经引入了全面的，从开始到结束的诱变能力。 这些变化对于促进新兴的基因组工程的发展是必要的 比如CRISPR。TTML团队与LCC成员合作， 修饰策略和基因分型测定，并为研究者提供确认的创始人。这些新 能力，除了我们的传统方法，创造转基因和ESC衍生突变株系，使 研究人员有机会采用总体方法，通过遗传修饰诱变， 设施。 TTML设施对于Lurie癌症中心的研究人员的研究计划至关重要， 小鼠模型来研究恶性肿瘤的潜在机制，并验证转化项目的靶点。它 提供必要的基础设施，使中心的研究人员能够获得转基因技术， 通常需要昂贵的显微注射设备和具有显微注射专业知识的熟练人员， 显微外科手术、胚胎操作、畜牧业、ES细胞培养和基因组工程。此外，本发明还提供了一种方法， TTML工作人员提供内部咨询和所有转基因相关技术的经常性指导，包括 CRISPR介导的基因组编辑系统，转基因和靶向载体设计，适当的筛选 策略，DNA纯化方法，建立者小鼠的繁殖和分析，合适的进口方法 小鼠品系和冷冻保存选择。由于TTML作为一种资源被纳入Lurie Cancer 中心在1995年，中心研究人员一直是利用TTML的NU教师的主要群体， 强调其在LCC整体研究使命中的关键作用。

ABSTRACT – TRANSGENIC AND TARGETED MUTAGENESIS LABORATORY The Transgenic and Targeted Mutagenesis Laboratory (TTML) is a Northwestern University-wide resource dedicated to the generation, import, recovery/rederivation, and cryopreservation of genetically engineered mice. TTML provides Lurie Cancer Center (LCC) members a broad range of services including generation of transgenic, gene edited, and targeted transgenic mouse models via microinjection; gene targeting/editing of embryonic stem (ES) cells; ES cell microinjection into blastocysts to create germline competent chimera; importation and recovery/rederivation of mouse lines from germplasm; and cryopreservation of mouse sperm and embryos. Since the last review, the core has introduced full-scale, start-to-finish mutagenesis capabilities. These changes were necessary to facilitate the development of newly emerging genome engineering technologies such as CRISPR. The TTML team works with LCC members to design customized genetic modification strategies and genotyping assays, and provides confirmed founders to investigators. These new capabilities, in addition to our traditional methods for creating transgenic and ESC derived mutant lines, enable investigators the opportunity to employ overarching approaches to genetic modification mutagenesis through the facility. The TTML facility is essential for the research programs of investigators of the Lurie Cancer Center who use mouse models to study mechanisms underlying malignancy and validate targets for translational projects. It provides the necessary infrastructure that allows the Center’s investigators access to transgenic technology that normally requires expensive microinjection equipment and skilled staff with expertise in microinjection, microsurgeries, embryo manipulation, animal husbandry, ES cell culture, and genome engineering. In addition, TTML staff provide in-house consultation and recurrent guidance on all transgenic-related technologies including CRISPR-mediated genome editing systems, transgenic and targeting vector design, appropriate screening strategies, DNA purification methods, breeding and analysis of founder mice, appropriate methods for importing mouse lines, and cryopreservation options. Since the inclusion of the TTML as a resource within the Lurie Cancer Center in 1995, Center investigators have consistently been the primary group of NU faculty utilizing the TTML, emphasizing its pivotal role in the overall research mission of the LCC.