斑马鱼二倍及单倍体胚胎干细胞培养和鉴定体系的研究

项目名称： 斑马鱼二倍及单倍体胚胎干细胞培养和鉴定体系的研究

项目编号： No.31271576

项目类型： 面上项目

立项/批准年度： 2013

项目学科： 生物科学

项目作者： 易梅生

作者单位： 中山大学

项目金额： 75万元

中文摘要： 胚胎干细胞（ES细胞）是生命科学研究的重要素材。然而，斑马鱼作为重要的模式脊椎动物，其ES细胞培养尚未成功，使以ES细胞为基础的研究工作难以开展。我们的前期研究表明，斑马鱼ES细胞培养的主要障碍在于培养条件不完善。本研究拟在前期工作的基础上，优化培养条件，建立有种系传递能力的斑马鱼ES细胞系，探索它们的生长特性、体内外分化潜能及有效的基因转移途径。斑马鱼ES细胞的培养成功将为基因修饰等研究提供有用的细胞平台。本研究还将在二倍体ES细胞研究的基础上，分离和培养雌核发育单倍体胚胎的囊胚细胞，建立斑马鱼单倍体ES细胞培养和鉴定体系。相比于二倍体ES细胞，单倍体ES细胞更适于遗传筛选和基因功能分析等研究，因为其基因组中每一基因只有一份，基因的改变可直接从表型上反映出来。该体系的建立，不仅可在理论上阐明斑马鱼单倍体基因组对ES细胞在体外生长发育的支持能力，而且还将提供一个更有效的斑马鱼基因研究工具。

中文关键词： 斑马鱼；细胞培养；胚胎干细胞；原始生殖细胞；抗病毒

英文摘要： Owing to their unique features, embryonic stem (ES) cells derived from early developing embryos provide a promising tool for experimental analyses of basic developmental processes such as pluripotency control and cell fate decision. As a model organism, zebrafish are widely used in basic studies in developmental biology and medicine. However, many effective ES cells-based methods and techniques currently used in other model organisms can not be applied in zebrafish duo to the lack of an excellent zebrafish ES cell culture system. Previous attempts of derivation of zebrafish ES cell long-term cultures were not successful. We consider that the failure was because of the less appropriate culture conditions which led to variations in chromosome number and loss of cell pluripotency. Our preliminary data suggest that it is possible to develop zebrafish long-term ES cell cultures in an appropriate culture system optimized by defining the components of medium. In this project, we propose to further improve the culture conditions and derive zebrafish ES cell cultures capable of germ line contribution. The features of cultivated ES cells are to be characterized by cytogenetics analysis, clonal growth, gene transfer experiment, ES cell marker expression analysis, embryoid body formation, intra- and inter- species chimera f

英文关键词： zebrafish；cell culture；embryonic stem cell；primordial germ cell；antivirus