萌发期大豆总蛋白提取及双向电泳体系的优化.

Sample preparation and electrophoresis conditions play crucial roles in proteomic technologies, yet they can be problematic. In this study, we compared three protein extraction methods, including TCA/actone method, phenol extraction and phenol/SDS extraction. We also optimized the conditions and established parameters of two-dimensional electrophoresis for the soybean seed during germination. The results showed that phenol/SDS extraction was the most suitable one. For 2-DE, 0.2mg samples were applied into IPG strips of pH4-7(Bio-Rad,17cm), subjected to IEF as follows: 250V for 1h, 500V for 1h, 3000V for 3h, 8000V for 1h, 10000V for 1h, and then 10000V until reaching 60000Vh. This study established a solid foundation for the proteomic analysis of soybean seed germination. [ABSTRACT FROM AUTHOR]

本文比较了TCA/丙酮法、酚抽提、苯酚/SDS法3种不同的蛋白提取方法, 并对双向电泳的一些条件进行了优化, 建立了适合萌发期大豆总蛋白的双向电泳体系, 即采用17cm、pH4-7的IPG胶条, 凝胶浓度为12%, 蛋白质上样量为0.2mg。IEF程序为：250V 1h,500V 1h,3000V 3h,8000V 1h,10000V 1h,10000V 60000Vh。本研究为萌发期大豆的蛋白组学研究提供了基础技术. [ABSTRACT FROM AUTHOR]