cspA冷休克启动子-重组人bFGF表达载体的构建及在大肠杆菌中可溶性表达研究  

Construction of cspA cold-shock promoter-recombinant human bFGF expression vector and its soluble expression in E. coli

作　　者：王振 邓超[2] 程咏梅 赵善成 付海田 陈敬华[1,3] 

机构地区：[1]江南大学药学院,无锡214122 [2]江南大学无锡医学院,无锡214122 [3]江南大学糖化学与生物技术教育部重点实验室,无锡214122

出　　处：《工业微生物》2015年第4期12-18,共7页Industrial Microbiology

基　　金：教育部新世纪优秀人才支持计划(No.NCET-10-0435);教育部博士点基金(No.20110093110008)资助

摘　　要：碱性成纤维生长因子b FGF具有刺激血管生成,促进血管内皮细胞和成纤维细胞增殖等生物活性,在组织修复中也起到重要作用。本研究运用逆转录聚合酶链式反应(RT-PCR),扩增出b FGF的编码序列,克隆至表达载体p ColdⅡ,构建csp A冷休克启动子调控的重组表达载体p Coldb FGF,转化E.coli BL21(DE3),在20℃低温诱导下b FGF几乎完全是可溶性表达,表达量占细胞总蛋白50%以上。纯化的b FGF能够促进NIH 3T3增殖,半数有效剂量ED50为1.33μg/L,具有较高的生物活性。Basic fibroblast growth factor possesses very important biological activities in vivo such as stimulating angio- genesis and promoting proliferation of vein endothelial cell and fibroblast, which plays a role in tissue repair. However, deficient source of nature bFGF and poor solubility and biological activity prevent its extensive use and research. In this paper, the coding sequence of bFGF was amplified by RT-PCR and cloned into the expression vector pCold Ⅱ containing cold-shock promoter cspA. Under low-temperature induction at 20 ℃, expression of bFGF was almost soluble with a more than 50% of the total cellular protein in E. coli BI21 （ DE3 ） transformed by pCold-bFGF. The purified bFGF had good biological activity which could effectively promote the proliferation of NIH 3ri3 and the EDs0 was 1.33 μg/L.

关 键 词：bFGF cspA冷休克启动子 E.COLI B1221(DE3) 可溶性表达 

分 类 号：Q78[生物学—分子生物学]