FISH EMBRYONAL STEM CELLS FOR DEVELOPMENT OF TRANSGENICS

用于转基因开发的鱼胚胎干细胞

• 批准号: 2154837
• 负责人: DAVID William BARNES
• 金额: $ 12.39万
• 依托单位: OREGON STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1992
• 资助国家: 美国
• 起止时间: 1992-09-30 至 1995-09-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2154837
• 关键词: alternatives to animals in research; biotechnology; disease /disorder model; embryonic stem cell; genetically modified animals; mitogens; tissue /cell culture; transfection; trout /salmon; zebrafish

Work with other systems utilizing pluripotent embryonal stem (ES) cell cultures for the formation of chimeric animals has succeeded in creating organisms in which foreign genes are expressed, as well as organisms in which endogenous genes have been altered. Dr. Barnes is interested in developing Zebra fish ES cell cultures for derivation of chimeric transgenic animals. He developed serum-free culture techniques allowing derivation of karyotypically normal late-stage mouse embryo cells that could not be derived in conventional serum-containing medium. Applying this approach to fish embryo cells, he found a mitogenic activity in trout embryo extract that was essential for the cultures and adapted transfection techniques to achieve expression of exogenous genes in fish cell cultures. He proposes to: (1) use the mitogenic activity in trout embryos to derive ES cell cultures and characterize the cells through transfection of exogenous genes, injection into developing embryos, and evaluation of contribution to transgenic chimeras; (2) apply this approach to haploid embryos to derive haploid ES cell cultures allowing alteration of endogenous genes.

与其他使用多能胚胎茎（ES）细胞的系统一起工作 形成嵌合动物的培养已经成功地创造了 表达外源基因的生物以及生物 哪些内源基因已改变。 巴恩斯博士对 开发斑马鱼ES细胞培养物以推导嵌合 转基因动物。 他开发了无血清的培养技术 核型在核型正常的晚期小鼠胚胎细胞的推导 无法在传统的含血清的培养基中得出。 申请 这种鱼类胚细胞的方法，他在鳟鱼中发现了有丝分裂活性 对于培养物和适应转染至关重要的胚胎提取物 在鱼细胞培养物中实现外源基因表达的技术。 他提出：（1）在鳟鱼胚胎中使用有丝分裂活性来得出 ES细胞培养并通过转染来表征细胞 外源基因，注射到发育的胚胎中，评估 对转基因嵌合体的贡献； （2）将此方法应用于单倍体 胚胎得出单倍体ES细胞培养物，允许改变 内源基因。