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目的:探讨细胞质膜及其蛋白质复合物分离与定性的方法。方法:结合差速离心法、密度梯度离心法、双水相法分离得到细胞质膜,并利用透射电镜和Western blot等实验手段验证分离所得细胞质膜的纯度。结果:本文采用了蓝绿温和凝胶电泳法对细胞质膜上的蛋白质复合物进行一维分离,得到23种蛋白质复合物;接着对23种蛋白质复合物进行第二相垂直电泳分离,分离出135种蛋白质;最后鉴定78种是质膜相关蛋白,其余57种来源内膜系统。结论:多种提纯方法的联用可以有效提高蛋白分离率与分离纯度,高纯度的质膜对质膜蛋白质组的研究具有重要意义。
Objective: To explore the method of separating and characterizing the plasma membrane and its protein complex. Methods: The plasma membrane was separated by differential centrifugation, density gradient centrifugation and aqueous two-phase method. The purity of the isolated plasma membrane was verified by transmission electron microscopy and Western blot. Results: In this study, 23 kinds of protein complexes were obtained by one-dimensional separation of the protein complexes on the plasma membrane by using blue-green mild gel electrophoresis. Then the 23 kinds of protein complexes were subjected to the second-phase vertical electrophoresis separation to separate 135 78 kinds of proteins were identified as plasma membrane-associated proteins and the remaining 57 were derived from the intima system. Conclusion: The combination of multiple purification methods can effectively improve the protein isolation rate and separation purity, and the high purity plasma membrane is of great significance to the research of plasma membrane proteome.