利用不同电泳技术对蝎毒素ITX蛋白组分分离的实验研究

【作者】 高峰； 曹俊娜； 齐伟； 高革； 高孟哲； 刘文第；

【机构】 郑州大学河南省医药科学研究院、医学实验中心； 河南中医学院； 新乡医学院测试中心；

【摘要】 目的:初步建立蝎毒素 ITX(Immunotoxin)蛋白组分的电泳分离方法和蛋白质组表达谱。方法:阴极电泳(T=18%,C=6%)对蝎毒进行分离,Bio-Rad Quantity-one 电泳软件分析条带;双向电泳技术:第一向为固相 pH 梯度等电聚焦,第二向电泳为垂直平板 SDS-PAGE(T=18%)。在以上过程中分别对上样方式、等电聚焦程序等条件进行优化控制,利用 PDQ7·1软件分析获得二维凝胶图像。结果:1、阴极电泳图谱显示蝎毒素 ITX 由7个蛋白组分组成:2、二维凝胶图谱显示蝎毒素 ITX 包含分子量相同(约为5800D)而等电点不同9种蛋白组分,其中酸性蛋白质4种,碱性蛋白质5种。结论:通过各种条件的优化, 初步建立了蝎毒素 ITX 的阴极电泳和双向电泳方法,获得了分辨率和重复性较好的阴极电泳图谱和二维凝胶图谱。更多还原

【Abstract】 Objective:With techniques of negative pole electrophoresis and 2-DE(Two-dimensional gel electrophoresis),the thesis analyzes the separation method and the proteome of scorpion toxin ITX(Immunotoxin).Method.By separating the scorpion toxin ITX with techniques of negative pole clectrophoresis(T=18%,C=6%),straps are analyzed with Bio-Rad Quantity-one:Ampholine electrophoresis of solid phase pH gradient,and the second approache electrophoresis is the vertical plate SDS-PAGE(T=18%).During the above process, the method of application and the process of isoelectrofocusing are optimized.Electrophoregram is analyzed with PDQuest7.1.Result:7 protein straps were visualized on negative pole electrophoregram,9 protein spots were visualized on 2-DE gel by Coomassie brilliant blue staining,in which molecular mass were about 5800D.And 4 proteins were abundant.Based on their pIs.,4 proteins were acidic and 5 proteins were distributed in the basic area.Conclusion: Negative pole electropboregram and 2-DE electrophoregram were gotten,which have great resolution and reproducibility.The separation techniques of negative pole electrophoresis and 2-DE of scorpion toxin ITX were set up with grading-up the conditions.更多还原