金鱼胚胎蛋白质双向凝胶电泳方法建立及优化  

Establishment and Optimization of Two-dimensional Gel Electrophoresis Condition for Goldfish Embryos Proteome

作　　者：喻小燕[1,2] 陈妍[2] 曹晶[2] 谢锦云[2] 

机构地区：[1]湖南信息职业技术学院,湖南长沙410200 [2]湖南师范大学生命科学学院,湖南长沙410081

出　　处：《金陵科技学院学报》2009年第2期91-96,共6页Journal of Jinling Institute of Technology

基　　金：国家自然科学基金项目(90408017)

摘　　要：以金鱼胚胎为材料，以不同样品制备方式、裂解液成分、固相pH梯度胶条的选择、凝胶浓度为侧重点，建立并优化金鱼胚胎总蛋白质组最佳双向凝胶电泳技术条件。研究表明，采用匀浆器匀浆含硫脲和NP-40裂解液提取蛋白，应用pH值为3～10的非线性IPG胶条和浓度14％的SDS-PAGE凝胶进行双向电泳，经优化电泳电流、时间等参数后，获得了重复性和分辫率理想的金鱼胚胎总蛋白质双向电泳图谱，为金鱼胚胎蛋白质组分析奠定了基础，为揭示金鱼单倍体发育异常的机制提供了技术保障。To develop and optimize a two-dimensional （2-DE） gel electrophoresis condition for gold{ish embryos proteome, total protein of goldfish embryos is extracted, and the optimal conditions for sample preparation and 2-DE are screened by comparing the following parameters： embryos broken methods, contents of lysis buffer, the concentration of gel, and linear or non-linear immobilized pH gradient gels. The results demonstrate that a satisfactory 2-DE map with good repetition and resolution is obtained and suitable conditions for 2-DE are established, with the conditions of the homogenate of goldfish embryos, adjusting IPG pH from 3 to 10 non-linear immobilized pH gradient gels and using 14 % SDS-PAGE gel as well as optimizing electrophoresis current, time and other parameters. The results paves a way for analysis of global protein profile in goldfish embryos, and lays a technological foundation for further study in the developmental mechanism of goldfish gynogenetic haploid embryos.

关 键 词：金鱼胚胎 蛋白质组 双向凝胶电泳 

分 类 号：Q51[生物学—生物化学] Q81