适用于黄瓜叶片、根系和果实总蛋白的双向电泳体系的建立  

Establishment of Two-dimensional Electrophoresis System Suitable for Cucumber Leave, Root and Fruit Proteins

作　　者：刘晓林[1] 杨学勇[1] 余宏军[1] 蒋卫杰[1] 赵文超[1] 王燕[1] 张晓孟 

机构地区：[1]中国农业科学院蔬菜花卉研究所,北京100081

出　　处：《基因组学与应用生物学》2016年第3期721-727,共7页Genomics and Applied Biology

基　　金：农业行业科技专项(201203001;201203095);国家大宗蔬菜产业技术体系(CARS-25-C-09);设施蔬菜协同创新中心;设施农业部园艺作物生物学与种质创制重点实验室共同资助

摘　　要：通过对上样量、分离胶浓度、等电聚焦参数三方面条件的优化,采用三氯乙酸/丙酮法提取黄瓜叶片总蛋白,初步建立了适用于黄瓜叶片总蛋白的双向电泳体系。进一步发现此优化条件结合蛋白的酚提取法,也适用于黄瓜根系和果实总蛋白的双向电泳。具体优化条件为：选用24 cm pH 4~7线性固相化pH梯度（immobilized pH gradient,IPG）胶条,上样量为800μg,分离胶浓度为12.5%,按聚焦程序Ⅱ聚焦,采用胶体考马斯亮蓝方法染色。采用该优化的体系可以同时进行黄瓜叶片、根系和果实总蛋白的双向电泳,并获得分辨率高、重复性好的双向电泳图谱。。A two-dimensional electrophoresis（2-DE） system for proteomic analysis of cucumber leaves was established using TCA/acetone precipitation method for total protein extract by optimizing the parameters including the loading amount of sample,concentrations of the second-dimensional SDS separation gel and isoelectric focusing conditions. We further found that this system was also suitable for cucumber roots and fruits using phenol extraction method coupled with ammonium acetate precipitation for total protein extract. The optimized system include the following steps： separating 800 μg protein samples with 24 cm linear pH 4~7 immobilized pH gradient（IPG） strips followed by isoelectric focusing program Ⅱ,and staining gels by colloidal Coomassie Brilliant Blue method after SDS-PAGE electrophoresis with 12.5% separation gel. Reproducible profiles with high resolution were obtained by this optimized 2-DE system for cucumber leaves,roots and fruits.

关 键 词：黄瓜 叶片 根系 果实 双向电泳体系 

分 类 号：Q946.1[生物学—植物学]