海滨木槿叶片蛋白质双向电泳体系的建立  

Establishment of 2-D PAGE System for Leaf Protein of Hibiscus hamabo

作　　者：李翠云[1,2,3] 姜彦成[1] 乔桂荣[2,3] 刘明英[2,3] 蒋晶[2,3] 卓仁英[2,3] 

机构地区：[1]新疆大学生命科学与技术学院,新疆乌鲁木齐830046 [2]中国林业科学研究院亚热带林业研究所,浙江富阳311400 [3]中国林业科学研究院林木遗传育种国家重点实验室,北京100091

出　　处：《西南林业大学学报（自然科学）》2012年第4期30-35,共6页Journal of Southwest Forestry University：Natural Sciences

基　　金：中央级公益性科研院所基本科研业务费专项资金项目(RISF6155)资助

摘　　要：为建立一套适合海滨木槿叶片蛋白质组学分析的双向电泳体系,以海滨木槿无性系叶片为材料,分别采用裂解液法、三氯乙酸-丙酮沉淀法、改良酚抽法分离纯化蛋白质,选用24 cm pH 3～10的IPG胶条,并对不同上样量、等电聚焦程序、平衡时间等电泳条件进行优化。结果表明:采用改良酚抽法提取蛋白质,上样量为每IPG胶条1 000μg,总聚焦功率90 kV.h,平衡时间15 min,胶体考马斯亮蓝染色,可得到蛋白质点数目多、分辨率高的双向电泳图谱。重复性试验结果显示,该体系具有很好的稳定性及可重复性,可满足海滨木槿蛋白质组学研究的要求。To establish a two-dimensional polyacrylamide gel electrophoresis （2-D PAGE） system for leaf pro- teomic analysis of Hibiscus hanmbo, three methods including lysis-buffer method, TCA-acetone precipitation meth- od, and the improved phenol extraction method were applied for leaf protein isolation and purification, and the key technologies of electrophoresis including the sample loading quantity, isoelectric focusing conditions, equilibrium time were optimized by taking 24 cm pH 3-10 immobilized pH gradient（IPG） strips. The results indicated that the reproducible 2-D gel with more spots and high resolution electrophoresis diagram could be obtained by using the im- proved phenol extraction method, with 1 000 μg of loading protein sample per IPG strip, 90 kV · h isoelectric focu- sing （IEF） time, 15 min of equilibrium time and staining with colloidal coomassie brilliant blue as the most suit- able treatment conditions. The results of repetitive experiments showed that this electrophoresis system was stable and reproducible, which could meet the demand for the proteomics research on H. hamabo.

关 键 词：海滨木槿 耐盐 蛋白质 双向电泳 

分 类 号：S718.46[农业科学—林学]