CORE--TRANSGENIC MOUSE/ES CELL CHIMERA FACILITY

核心——转基因小鼠/ES细胞嵌合体设施

• 批准号: 6315259
• 负责人: ALEXANDRA L. JOYNER
• 金额: $ 13.42万
• 依托单位: NEW YORK UNIVERSITY SCHOOL OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-05-19 至 2002-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6315259
• 关键词: animal colony; biomedical facility; embryonic stem cell; genetically modified animals; laboratory mouse; neoplasm /cancer; neoplasm /cancer genetics

The transgenic Mouse/ES Cell Chimeras Facility provides four basic services: production of transgenic (Tg) mice by zygote injection, production of embryonic stem (ES) cell chimeric animals, rederivation of mouse strains and isolation of gene-targeted ES cells following homologous recombination. For production of transgenic mice, users supply DNA constructs suitable for pro-nuclear injection into fertilized mouse eggs. Transgenic mice are produced in outbred Swiss Webster mice, inbred FVB/N mice or mice of defined genotype. Potential founder animals are provided to the research investigator at weaning for DNA analysis by Southern blotting for genotyping. ES cell chimeras as produced by morulae aggregation using the Swiss Webster strain as a host or by injection of ES cells into C57BL/6 blastocysts. Chimeras are provided to the research investigator at weaning for test breeding to determine germline transmission of the mutation. Aggregation chimeras between two strains of mice can also be made through the TgESC facility. A recent service being offered involved the new technique of making chimeras with tetraploid morulae to produce embryos that are 100% ES cell-derived. To enable investigators to house pathogen-free animals in the SPF animal quarters in Skirball, rederivation of mouse strains is also offered. For isolation of gene-targeted ES cells, users of the resource supply DNA targeting constructs prepared in consultation with the facility and a confirmed genotyping assay. Constructs are electroporated into ES cells prepared in the facility, and isolated clones are scored using the genotyping assay. Positive homologous recombinants are transferred to the chimera unit for transmission of the mutant allele into mice. In addition, the facility provides researchers with a wide array of services related to the production of transgenic and knockout ES cells and chimeric mice such as supply of superovulated egg donors, pseudopregnant females, training in mouse husbandry and consultation in transgenic/ES cell approaches and vector design.

转基因小鼠/ES细胞嵌合体设施提供了四个基本的 服务：通过受精卵注射生产转基因（Tg）小鼠， 胚胎干（ES）细胞嵌合动物的产生， 小鼠品系和基因靶向ES细胞的分离， 同源重组为了生产转基因小鼠，用户 提供适合于前核注射入受精卵DNA构建体 老鼠卵转基因小鼠是在远系繁殖的瑞士韦伯斯特小鼠中产生的， 近交系FVB/N小鼠或确定基因型的小鼠。潜在创始动物 在断奶时提供给研究者进行DNA分析， Southern印迹法进行基因分型。ES细胞嵌合体， 使用瑞士韦伯斯特菌株作为宿主或通过 将ES细胞注射到C57 BL/6囊胚中。嵌合体被提供给 研究人员在断奶时进行试验育种， 突变的生殖细胞传播。两个之间的聚集嵌合体 小鼠品系也可以通过TgESC设施制备。最近的一 提供的服务涉及制造嵌合体的新技术， 四倍体桑椹胚以产生100%ES细胞衍生的胚胎。到 使研究者能够在SPF动物中饲养无病原体的动物 在Skirball的宿舍，还提供了小鼠品系的再衍生。为 分离基因靶向的ES细胞，用户的资源供应DNA 与设施协商准备的目标结构， 确认的基因分型试验。将构建体电穿孔到ES细胞中 在设施中制备，并使用 基因分型测定。将阳性同源重组体转移至 用于将突变等位基因传递到小鼠中的嵌合体单位。在 此外，该设施还为研究人员提供了广泛的 转基因和基因敲除ES细胞生产相关服务 和嵌合小鼠如超数排卵卵子供体的供应， 假孕雌性，小鼠饲养培训和咨询 转基因/ES细胞方法和载体设计。