【摘要】 以银杏小孢子叶球为材料,比较了不同的提取的方法、蛋白质裂解液组成、pH值范围以及不同质量浓度SDS-PAGE凝胶对双向电泳的影响。结果表明:TCA/丙酮/酚法比TCA/丙酮法更适合于银杏小孢子叶球蛋白的提取;含有硫脲的蛋白质裂解液Ⅱ,可显著提高蛋白质的溶解性,在2-DE图谱上可分辨出748个蛋白质点,比用蛋白质裂解液Ⅰ提取的蛋白点数多620个。采用pH3-10的胶条分离时,蛋白点集中在pH=4～8,使用pH=4～7的胶条,图谱胶点重叠少,也较清晰,更利于低丰度蛋白的检出。用TCA/丙酮/酚提取蛋白,并以蛋白质裂解液Ⅱ溶解蛋白,采用pH=4～7线性17 cm的IPG胶条1,2%SDS-PAGE凝胶进行双向凝胶电泳,银杏小孢子叶球蛋白质能得到较好的分离,图谱清晰。更多还原

【Abstract】 The effects of different protein extraction methods,lysis buffers,pH ranges of IPG strip and different gel concentrations on two-dimensional gel electrophoresis were studied by using male flower buds of Ginkgo biloba as experimental materials.Result shows that,compared TCA-acetone-phenol protocol with TCA-acetone protocol,the former is more suitable.The protein solubility of the samples dissolved in lysis bufferⅡcontaining thiourea was greatly enhanced,and about 748 protein spots were detected on 2-DE gels,which was 620 more than that in lysis buffer I.More spots were assembled from pH 4 to pH 8.Replaced pH 3-10 strips with pH 4-7 strips,the spots were shown in a clearer way.When the proteins were extracted by the TCA-acetone-phenol method,dissolved with the lysis buffer II,with 17cm,pH 4-7 IPG strip,and 12% concentrations of the SDS-PAGE gel,they were better separated.更多还原