红豆杉细胞蛋白质双向电泳样品制备方法的比较

Comparison of Protein Sample Preparation Methods from Taxus Cell for Two-dimensional Electrophoresis

[目的]探索适用于红豆杉愈伤蛋白质组学研究的样品制备方法。[方法]以红豆杉愈伤组织为材料,分别采用三氯乙酸(TCA)/丙酮沉淀法、Tris-HCl法和酚-甲醇/醋酸铵沉淀法提取其中的总蛋白,通过双向电泳(2DE)检测3种方法对总蛋白的提取效果。[结果]3种方法提取的蛋白样品2DE图谱显示的蛋白点数量均较多。TCA/丙酮沉淀法对蛋白质的提取率较高,2DE图像清晰,条纹少,拖带现象轻;Tris-HCl法提取的蛋白样品2DE图谱个别区域出现较轻的云片状聚集,并有轻微拖带现象;酚-甲醇/醋酸铵沉淀法提取蛋白样品的2DE图谱有条纹和拖带现象。[结论]TCA/丙酮沉淀法提取的蛋白质质量较高,所得图谱背景清晰,蛋白质信息量大。

[ Objective ] The aim was to study protein sample preparation method that suitable for proteomics research of taxus callus. [ Method ] With taxus callus as the materials, its total protein was extracted by trichioroaeetic acid （TCA）/acetone precipitation, Tris-HCl and phenol-methanol/ammonium acetate precipitation methods, the extraction effects of the 3 methods on total protein were detected by two-dimensional electrophoresis （2DE）. [ Result] Protein spots on 2DE map of total protein extracted by 3 methods were all more. The extraction rate of （TCA）/acetone precipitation method on protein was high, and its 2DE map was clear with less fringe and light entrainment phenomena. Some regions on 2DE map of total protein extracted by Tris-HCl method appeared slight cloud and flaky aggregation with slight entrainment phenomena. 2DE map of total protein extracted by phenol- methanol/ammonium acetate precipitation method had fringe and entrainment phenomena. [ Conclusion] The quality of protein extracted by （TCA）/acetone precipitation method was better, and the background of its 2DE map was clear with large-scale protein information.