摘要
通过优化诱导表达条件,实现抗镰刀菌单链抗体在大肠杆菌周质中高效可溶性表达。将抗镰刀菌单链抗体FvSG7转化到大肠杆菌XL1-Blue中,确定诱导表达培养基,在不同的诱导温度、诱导剂IPTG的浓度和诱导时间条件下培养重组大肠杆菌,通过Western杂交和ELISA检测分析单链抗体的可溶性表达情况以及抗体的活性。重组大肠杆菌培养至OD600nm为0.5时加入终浓度为0.1 mmol/L IPTG,25℃诱导表达2 h可获得最大量的可溶性单链抗体。通过对诱导温度、IPTG浓度和诱导时间等表达条件的优化,可以显著提高Fv SG7抗体在大肠杆菌XL1-Blue周质中的表达量。
The objectives of the work is to optimize the conditions for inducing expression, and obtain the soluble and high-yield expression of a Fusarium-specific single-chain variable fragment（scFv）antibody in the periplasmic space of Escherichia coli XL1-Blue. The recombinant plasmid containing a Fusarium-specific scFv antibody FvSG7 was transferred into E. coli XL1-Blue. After the culture medium for inducing selected, the soluble expression level and activity of FvSG7 antibody were analyzed by Western blot and ELISA detection for studying the influence of temperature, IPTG concentration and induction time on expression level. The maximum productivity of soluble FvSG7 antibody was obtained after induction at 25℃for 2 h, with a final concentration of 0.1 mmol/L β-D-thiogalactopyranoside（IPTG）and the cultured bacteria growing to the OD600 value of 0.5. In conclusion, the soluble expression of FvSG7 antibody in the periplasm of E. coli XL1-Blue increased significantly by optimizing the expression’s conditions of induction temperature, IPTG concentration and induction time.
出处
《生物技术通报》
CAS
CSCD
北大核心
2015年第9期238-243,共6页
Biotechnology Bulletin
基金
国家重点基础研究发展计划项目(2013CB127801)
国家自然科学基金项目(31201475)
关键词
单链抗体
可溶性表达
大肠杆菌
优化
single-chain variable fragment antibody
soluble expression
Escherichia coli
optimization
