抗克伦特罗单链抗体-碱性磷酸酶融合蛋白表达条件的初步研究

Expression conditions of the recombinant anticlenbuterol single-chain Fv antibody-alkaline phosphatase fusion protein

初步研究抗克伦特罗单链抗体-碱性磷酸酶融合蛋白(CBLscFv-AP)的表达条件,提高CBLscFv-AP融合蛋白的表达量。采用液体发酵法培养抗克伦特罗单链抗体-碱性磷酸酶融合蛋白(CBLscFv-AP)基因工程菌,研究不同宿主菌、初始pH、诱导时期、诱导时间和培养基对CBLscFv-AP融合蛋白表达量的影响以及重组质粒的稳定性。选用E·coliBL21(DE3)pLysS为宿主菌,在初始pH为7·4的LB培养基中培养至A600nm为0·8～0·9时,诱导表达8h,为CBLscFv-AP融合蛋白最佳表达条件。而且重组质粒pBV220-scFv-phoA具有较好的结构稳定性。在该表达条件下,提高了CBLscFv-AP融合蛋白的表达量,为进行大规模发酵生产CBLscFv-AP融合蛋白奠定基础。

The expression conditions of recombinant E.coli strain BL21（DE3）pLysS expressing recombinant anticlenbuterol single-chain Fv antibody-alkaline phosphatase fusion protein were studied.The E.coli host cells,initial pH of culture medium,induction period,induction time and culture medium were optimized,and the stability of plasmid pBV220-scFv-phoA was also investigated.The expression yield of the CBLscFv-AP fusion protein was enhanced,when recombinant E.coli strain BL21（DE3）pLysS harboring plasmid pBV220-scFv-phoA was induced for 8 hours after the bacterial density A600nm reached 0.8 ~ 0.9 in LB medium.The plasmid pBV220-scFv-phoA in E.coli strain BL21（DE3）pLysS expressed CBLscFv-AP fusion protein stability,after being cultured to 100 generations.The developed expression conditions could increase the expression yield of CBLscFv-AP fusion protein,and were suitable for the production of CBLscFv-AP fusion protein in a large scale.