大豆叶片蛋白质双向电泳技术的改良  

Improvement of Two-dimension Electrophoresis for Soybean Leaves Total Protein

作　　者：顾宪鹏[1] 罗秋兰[1] 高继国[1] 李文滨[1] 

机构地区：[1]东北农业大学大豆研究所教育部大豆生物学重点实验室,黑龙江哈尔滨150030

出　　处：《大豆科学》2011年第4期663-667,共5页Soybean Science

基　　金：国家自然科学基金面上资助项目(30971810)

摘　　要：以大豆叶片为材料,比较分析了3种蛋白质的提取方法,讨论了等电聚焦时间和染色方法等关键因素对双向电泳的影响,同时选择不同的IPG胶条,设定了第1向等电聚焦和第2向SDS-PAGE电泳的电泳程序及参数。结果表明：采用TCA-丙酮沉淀法提取蛋白质,选择pH 4~7（17 cm）IPG胶条,适当延长和改变等电聚焦的第2步和第3步的时间和参数,并且用考马斯亮蓝和银染复合染色法对12%的凝胶进行染色,能够获得清晰蛋白点的双向电泳图谱,说明优化出的双向电泳技术更加适合于大豆叶片总蛋白质分离。To improve the two-dimensional polyacrylamide gel electrophoresis（2D-PAGE）technology for separating leaf proteins in soybean,we compared three soybean leaf protein isolation methods,discussed the influence of isoelectric focusing time and staining,chose the different IPG strips to set the electrophoresis program and parameters of the first isoelectric focusing and the second sodium dodecyl sulfate polyacrylamide gel electropheresis（SDS-PAGE）.When chose the TCA-acetone precipitation method to extract protein,selected pH 4-7（17 cm）IPG strips,prolonged the isoelectric focusing time of the second and the third step,modified the isoelectric focusing parameters,and combined the coomassie brilliant blue staining and silver staining for 12% gelatin,we obtained clear soybean protein 2D-PAGE map.Results suggest the optimized method is suitable for separating total protein in soybean leaf.

关 键 词：蛋白质提取 SDS-PAGE 双向电泳 蛋白质组学 

分 类 号：S565.1[农业科学—作物学]