适用于毛白杨芽双向电泳分析的蛋白质提取方法

Optimal protein extracting methods of Populus tomentosa buds for two-dimensional gel electrophoresis

本实验以毛白杨芽为实验材料,采用Tris-酚法、三氯乙酸-丙酮法和三氯乙酸-丙酮+Tris-酚法对其蛋白质提取方法进行比较,采用双向电泳分离蛋白质,通过PDQuest软件和质谱对这3种提取方法得到的双向电泳胶进行分析。结果显示,三氯乙酸-丙酮法得到的蛋白点数量为231±4,得到的鲜样中的总蛋白质含量为(3.57±0.618)mg/g,均高于其他两种提取方法,并且所得到的双向电泳胶更清晰,为毛白杨芽蛋白质提取的最适方法。

Three protein extracting methods （Tris-phenol method, TCA/acetone method and TCA/acetone ± Tris-phenol method） were compared with the materials of P. tomentosa buds and described in this report. To determine which protein extracting method is most suitable for P. tomentosa buds, two-dimensional electrophoresis was applied to the separation of proteins, PDQuest software and mass spectrometry were used to characterize the protein spots from 2D-gel. Among the three protein extracting methods, TCA/ acetone method was the most suitable one for P. tomentosa buds and yielded （3.57 ± 0. 618） mg/g total protein amount in fresh sample, 231 ± 4 protein spots on the 2D-Gel and the clearest 2D-Gel images, which were higher in total amount, more in spot number and clearer in 2D-Gel image than other two methods.