摘要
将ScFv基因片段与pHOG21载体连接后,转化至受体菌XL1-Blue中,得到重组菌株XL1-Blue（pHOG2E3）。随后研究了培养基中无机盐成分、温度、诱导时间、IPTG和蔗糖浓度对ScFv基因表达的影响。经SDS-PAGE分析表明,重组菌株XL1-Blue（pHOG2E3）在LB培养基中加入0.5mmol/L IPTG和 0.4 mol/L蔗糖,37℃诱导6h,其目的蛋白的表达量较高,表达的ScFv蛋白主要以包含体的形式存在,分子量为31,000D,在重组菌株的培养上清和菌体裂解上清液中,通过ELISA法也检测到了ScFv的存在,且具有中和α毒素的活性。
The ScFv gene containing Nco I and Bam HI was amplified by PCR, and inserted into the prokaryotic expression vector pHOG21 with Nco I and Bam HI digestion. After screening, high-level expression recombinant XL1-Blue (pHOG2E3) were obtained, and the different expression levels of ScFv were detected by SDS-PAGE and EUSA. When the strain was induced by 0.5 mmol/LIPTG and 0.4mol/L sucrose in the culture medium LB at 37℃ for 6 hours, the high level production of ScFv protein was obtained. The molecular weight of the expressed ScFv is 31, 000 D. The ScFv was mainly in the form of inclusion body, but it could also be in the culture medium and soluble periplasmic content. Above all, the ScFv protein could neutralize the phospholipase C activities of alpha-toxin of Gostridium perfrigens type A.
出处
《微生物学通报》
CAS
CSCD
北大核心
2002年第2期8-13,共6页
Microbiology China
关键词
产气荚膜梭菌a毒素
单链抗体
基因表达
中和活性
Alpha- toxin of Clostridium perfringens, ScFv, Gene expression, Neutralization effect
