摘要
目的采用双向电泳分析水蛭(Hirudo)酒炙前后差异蛋白表达。方法分别采用饱和硫酸铵沉淀法、三氯乙酸/丙酮沉淀法、直接裂解液法提取生水蛭和酒炙水蛭蛋白,十二烷基硫酸钠-丙烯酰胺凝胶电泳(SDS-PAGE)对这3种方法进行筛选,建立双向电泳体系。分析水蛭酒炙前后蛋白表达谱,寻找相关差异蛋白。结果直接裂解液法所得条带数量最多,分布连贯清晰。水蛭酒炙前后蛋白含有量有显著性差异(P<0.01),共发现19个差异蛋白点,其中下调蛋白8个,上调蛋白11个。结论双向电泳所得差异蛋白可作为蛋白标志物以规范水蛭炮制工艺,并确保其稳定性。
AIM To analyze the differential protein expressions in Hirudo before and after stir-frying with wine by two-dimensional eletrophoresis. METHODS Saturated ammonium sulfate precipitation,trichloroacetic acid /acetone precipitation and direct pyrolysis liquid methods were applied to extracting proteins from raw and wine-fried Hirudo,respectively. Then these three methods were screened by sodium dodecyl sulfate polyacrylamide gel electrophoresis（ SDS-PAGE）,and the two-dimensional eletrophoresis system was established. The protein expression profiles of Hirudo before and after stir-frying with wine were analyzed to find out related differential proteins.RESULTS The most number of strips was obtained by direct pyrolysis liquid method,whose distribution was coherent and clear. The contents of proteins extracted from Hirudo before and after stir-frying with wine showed obvious differences（ P〈 0. 01）. A total of nineteen differential protein points were found,eight of which were downregulated proteins,eleven of which were up-regulated proteins. CONCLUSION The differential proteins obtained by two-dimensional eletrophoresis can be taken as protein makers for regulating preparing process of Hirudo and ensuring its stability.
出处
《中成药》
CAS
CSCD
北大核心
2017年第2期360-365,共6页
Chinese Traditional Patent Medicine
基金
国家自然科学基金(81673596)
北京市教育委员会科技计划项目(KM201610025011)
首都医科大学科研基金项目(2015ZR26)
关键词
水蛭
酒炙
差异蛋白
SDS-PAGE
双向电泳
Hirudo
stir-frying with wine
differential proteins
SDS-PAGE
two-dimensional eletrophoresis
