利用双向电泳技术分离酿酒酵母全蛋白质,并比较了不同样品制备方法、不同凝胶参数,以及不同染色方法对电泳结果的影响.经过各个环节的条件优化,最终获得了无明显横纵条纹,背景清晰,分辨率较高,重复性较好的酿酒酵母全蛋白质双向电泳图谱.经PD-Quest软件分析,所得图谱约分离到1 000个蛋白质点,该双向电泳图谱的构建为后续氮代谢研究奠定了基础.
Total proteins of Saccharomyces cerevisiae was extracted and separated by two- dimensional electrophoresis (2-DE) in this study. Different sample preparation methods,different parameters of the gel and different staining methods were compared for the optimization of 2-DE. Finally,2D-gel without obvious vertical and horizontal stripes,with low background,high resolution and good reproducibility was obtained. Analyzed by the PD-Quest software,about l 000 protein spots were isolated from the 2-DE protein map. The construction of 2-DE protein map provide a firm basis for the follow-up nitrogen metabolism study.