产普鲁兰酶重组大肠杆菌质粒稳定性的研究  

Study on plasmid stability of recombinant Escherichia coli producing pullulanase

作　　者：翟成一 徐岩[1] 聂尧[1] 穆晓清[1] 

机构地区：[1]工业生物技术教育部重点实验室江南大学酿酒科学与酶技术中心,江苏无锡214122

出　　处：《工业微生物》2015年第2期13-20,共8页Industrial Microbiology

基　　金：国家高技术研究发展计划(863计划);食品酶及应用(2012AA022207);江苏高校优势学科建设工程资助项目

摘　　要：通过对产普鲁兰酶的重组大肠杆菌E.coli BL21(DE3)/p ET28a-s-pul菌株在发酵过程中质粒稳定性和普鲁兰酶生成量的考察,发现不同宿主对质粒稳定性及酶活性有重要影响。本文利用E.coli BL21(DE3)p Lys S菌株为宿主,构建重组菌E.coli BL21(DE3)p Lys S/p ET28a-s-pul,通过控制外源蛋白的本底表达,提高了重组菌株的质粒稳定性。优化发酵培养基和发酵条件以后,重组菌产普鲁兰酶能力由480 U/m L提高至627 U/m L,增幅为30.6%。研究结果认为,严格控制外源蛋白的本底表达,是改善重组菌稳定性的重要方法之一。It was found that different host bacteria would seriously affect the plasmid stability and pullulanase activity by investigating the recombinant E. coli BL21 （DE3）/pET28a-s-pul producing pullulanase. In this study,a recombinant E. coli BL21（DE3）pLysS/pET28a-s-pul was constructed with a new strain named E. coli BL21（DE3）pLysS,which suc-cessfully controlled the basal expression and improved the plasmid stability. After optimizing the medium and fermentation conditions,the pullulanase activity was enhanced from 480 U/mL to 627 U/mL with an increase of 30. 6%. The results of this research illustrated that strictly controlling the basal expression of the foreign protein was one of the important methods to improve the stability of recombinant bacteria.

关 键 词：普鲁兰酶 大肠杆菌 质粒稳定性 本底表达 发酵优化 

分 类 号：TQ920.1[轻工技术与工程—发酵工程]