苦瓜种子蛋白质组双向电泳技术条件的优化

摘要

通过对苦瓜种子蛋白质样品的提取方法、等电聚焦参数、SDS-PAGE分离胶浓度以及胶条pH范围的优化筛选,建立了适合苦瓜种子蛋白组分析的双向电泳体系.结果表明:使用TCA-丙酮提取方法提取苦瓜种子蛋白,更适用于双向电泳分析,结合使用pH=5~8 IPG胶条以及优化的聚焦程序,能获得分辨率较高、蛋白点清晰、重复性好的2-DE图谱.经银染显色,可检测约660个蛋白点,主要分布在p H=5~8;该体系适合用于分析苦瓜种子的蛋白质组.%Through optimization and screening of two-dimensional electrophoresis sample preparation methods of seeds protein of Momordica charantia ,the procedure of isoelectric focus ,concentration of the second-dimensional SDS separation gel and the pH range of solidifying strips ,electrophoresis sys-tem that was suitable to seed protein of Momordica charantia was structured .The results showed that highly pure seed protein can be obtained with trichloroacetic acid (TCA)-acetone method ,2-DE images of protein of high resolution ratio and good repetitiveness can be got with optimized isoelectric focus procedure .About 660 clear protein spots were detected by two dimensional electrophoresis (2-DE) and silver staining with isoelectric points at pH 5-8 .