安徽农业大学学报2018，Vol.45Issue(6)：1102-1106,5.DOI:10.13610/j.cnki.1672-352x.20190102.023
OjCHS原核表达载体的构建及重组蛋白的纯化
Prokaryotic expression vector construction and recombinant proteins purification of OjCHS
摘要
Abstract
Chalcone synthase (CHS) is the first and rate-limiting enzyme in the biosynthesis of flavonoids, and also plays important role during the biosynthesis and accumulation of flavonoids. In this study, we inserted OjCHS into pET-28 a vector to obtain recombinant pET28-CHS on the basis of the sequence of OjCHS. Then the recombinant plasmid was transfected into E.coli competent cell BL21 using a freeze-thaw method for the expression of recombinant proteins. Subsequently, the optimally induced expression condition was confirmed including IPTG concentration, inducing temperature and inducing time. Then a large number of soluble recombinant proteins were obtained through eluting with Ni column purification, further dialysis and concentration. The results showed that recombinant pET28-CHS vector was constructed and the optimally induced expression condition was 25 ℃ for 12 h by 0.45 mmol·L-1 IPTG induction. Finally, we obtained abundant soluble recombinant protein which was consistent with the expected size, which would lay a foundation for the further study of the biological functions of OjCHS.关键词
日本蛇根草/查尔酮合酶基因/原核表达载体/蛋白表达与纯化Key words
Ophiorrhiza japonica/chalcone synthase gene/prokaryotic expression vector/expression and purification of protein分类
生物科学引用本文复制引用
孙威,林建,申欢,彭贵,鞠志刚,马玲,乙引..OjCHS原核表达载体的构建及重组蛋白的纯化[J].安徽农业大学学报,2018,45(6):1102-1106,5.基金项目
国家自然科学基金(31760076) （31760076）
贵州省教育厅青年科技人才成长项目(黔教合KY字[2017]122) （黔教合KY字[2017]122）
贵州省联合基金项目(黔科合LH字[2016]7211号,黔科合LH字[2017]7358号) （黔科合LH字[2016]7211号,黔科合LH字[2017]7358号）
贵州省中医药管理局中医药、民族医药科学技术研究课题(S20160829000) （S20160829000）
贵州省重点实验室建设项目(黔科合计Z字[2011]4005)共同资助 （黔科合计Z字[2011]4005）
