橡胶草叶片蛋白质提取方法的选择及双向电泳体系的优化  

Isolation method of total protein from taraxacum kok-saghyz rodin and optimization of two-dimensional electrophoresis system

作　　者：李忠晴 柴武慧 张云川 黄叶红 李金环 鲁培鑫 闫洁[1] Li Zhongqing;Chai Wuhui;Zhang Yunchuan;Huang Yehong;Li Jinghuan;Lu Peixing;Yan jie(College of Life Science,Shihezi University,Shihezi,Xinjiang 832003,China)

机构地区：[1]石河子大学生命科学学院,新疆石河子832000

出　　处：《石河子大学学报（自然科学版）》2018年第5期609-616,共8页Journal of Shihezi University(Natural Science)

基　　金：国家自然科学基金项目(31360060)

摘　　要：目的为了建立起橡胶草叶片组织的蛋白双向电泳体系。方法采用5种不同方法提取橡胶草叶片蛋白,并通过蛋白提取率和单向SDS-PAGE电泳的比较,筛选出改良TCA-丙酮沉淀法、酚-乙酸铵沉淀法、Tris浸提法3种方法进行2-DE电泳。结果表明选择改良TCA-丙酮法为最合适的蛋白质提取方法。为进一步优化体系,比较了不同染色方法、上样量和固相p H梯度预制胶条(IPG)的p H范围对双向电泳结果的影响,电泳结果表明使用17 cm p H 3-10的IPG胶条、上样量为1200μg、考马斯亮蓝染色法能分离得到更多的蛋白点,是橡胶草叶片双向电泳体系的最优条件。结论本研究初步建立了橡胶草叶片蛋白质双向电泳体系,为深入研究橡胶草蛋白质组学提供依据。Objective In order to establish an appropriate two-dimension gel electrophoresis(2-DE) system for Taraxacum kok-saghyz(TKS) Rodin, five methods were used to extract the leaf protein. M ethods Based on the extraction rate and SDS-PAGE,the improved TCA-acetone method, phenol extraction-ammonium acetate precipitation method and Tris extraction were chosen for2-DE electrophoresis. R esults The results showed that the improved TCA-acetone method was the most suitable protein extraction method. In order to optimize the system,the loading quantity of sample, dyeing methods and strip conditions were compared and analyzed. The way of 1200 μg protein sample, 17 cm(linear, pH 3-10) strip, Coomassie brilliant blue was chosen as the best conditions for 2-DE. Conclusion This study established a preliminary system for TKS, and provided a theoretical basis for the proteomics research of TKS.

关 键 词：橡胶草 蛋白质 双向电泳 

分 类 号：Q946.1[生物学—植物学]