利用溶氧调控型启动子Pvgb构建产surfactin的重组枯草芽孢杆菌  

Constructing recombinant Bacillus subtilis producing surfactin using aeration-inducible promoter Pvgb

作　　者：周大袁 林佳辉 李霜[1] ZHOU Dayuan;LIN Jiahui;LI Shuang(College of Biotechnology and Pharmaceutical Engineering,Nanjing Tech University,Nanjing 211800,China)

机构地区：[1]南京工业大学生物与制药工程学院,江苏南京211800

出　　处：《生物加工过程》2020年第6期690-695,共6页Chinese Journal of Bioprocess Engineering

基　　金：国家自然科学基金(21576133);江苏省先进生物制造创新中心自主研发课题(XTD1824)。

摘　　要：表面活性素(surfactin)在石油开采、环境修复及医药卫生等行业应用广泛,但在生产过程中存在产生大量泡沫的难题。本文中,笔者尝试通过溶氧水平来调控surfactin的合成。溶氧诱导型启动子Pvgb使得目的基因受到溶氧水平的表达调控。以绿色荧光蛋白GFP为报告基因,考察Pvgb及其串联多拷贝启动子在枯草芽孢杆菌168菌株中的转录水平,结果发现,Pvgb启动子为组成型弱启动子,对溶氧水平的响应强度并不显著,Pvgb的拷贝数对表达水平具有"剂量效应",增加拷贝数,表达量增加。将不同拷贝数的Pvgb启动子以及强启动子P43分别构建回补Sfp功能蛋白的可产surfactin的B.subtilis168重组菌株,结果表明,Sfp功能蛋白的表达强度不是影响surfactin合成量的关键因素,溶氧供给水平是影响surfactin合成的关键因素。Surfactin is widely used in petroleum,exploitation,environmental restoration,medicine and health and other industries.However,there are a lot of bubbles in the production process of surfactin.In the study,we try to regulate the synthesis of surfactin by the level of dissolved oxygen.Aeration-inducible promoter Pvgb enables target genes to be regulated by the level of dissolved oxygen.The transcriptional level of Pvgb and its serial multiple copies in Bacillus subtilis 168 were investigated by using the green fluorescent protein(GFP)as the reporter gene.Pvgb was a weak promoter,and the response intensity to the level of dissolved oxygen was not significant.The level of the copy number of Pvgb had a"dose effect",and the amount of expression increased with increasing the number of copies.The Pvgb with different copy number and the strong promoter P43 were constructed to reconstruct the recombinant B.subtilis168 producing surfactin by supplementing of Sfp functional protein.The expression intensity of Sfp functional protein was not the key factor affecting the surfactin synthesis,and the level of dissolved oxygen was crucial to surfactin production.

关 键 词：表面活性素 Pvgb 溶氧诱导型启动子 溶氧调控 枯草芽孢杆菌 

分 类 号：Q812[生物学—生物工程]