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通过对不同裂解液配方、等电聚焦程序和SDS-PAGE方式等的对比实验,建立和优化了大弹涂鱼肝脏蛋白质组双向电泳的相关技术体系。结果显示采用裂解液中添加Tris和TBP,聚焦时适当延长除盐时间,提高聚焦电压和功率(伏-小时),能显著提高双向电泳图谱的分辨率,MSOG程序进行双向电泳(Multi-strips on One Gel MSOG),提高了双向凝胶的匹配率及有效性,降低了人为修饰点的增加,匹配率高达90%。通过相关条件优化提高了大弹涂鱼肝脏蛋白双向电泳图谱的分辨率,为后续大弹涂鱼的毒理蛋白质组学研究提供技术保障。
Through the comparative experiments of different lysate formulas, isoelectric focusing programs and SDS-PAGE methods, the related technology system of two-dimensional gel electrophoresis of liver proteome of Larvae Sinensis was established and optimized. The results showed that the addition of Tris and TBP to the lysis buffer could prolong the desalting time and increase the focusing voltage and power (V-hour) when focusing, which could significantly improve the resolution of two-dimensional gel electrophoresis. The MSOG program performed two-dimensional electrophoresis One Gel MSOG) improves the match rate and effectiveness of two-way gels and reduces the increase of artificial modification points with a matching rate of up to 90%. Through the optimization of the relevant conditions, the resolution of the two dimensional gel electrophoresis map of the liver was increased, which provided technical support for the toxicological proteomics study of the next bighead fish.