重组戊型肝炎疫苗抗原工程菌发酵工艺的优化  

Optimization of fermentation process of recombinant E.coli for expression of hepatitis E vaccine antigen

作　　者：徐军[1] 曹玉锋[1] 时成波[1] 李铮[1] 

机构地区：[1]长春生物制品研究所有限责任公司生物技术研究室,长春130062

出　　处：《中国生物制品学杂志》2012年第12期1697-1701,共5页Chinese Journal of Biologicals

基　　金：吉林省科技发展项目(20080940);吉林省产业技术研究与开发项目(2009003)

摘　　要：目的优化重组戊型肝炎疫苗抗原工程菌的发酵工艺,为其规模化生产奠定基础。方法以目的蛋白的表达量及菌体浓度作为综合评价指标,对重组戊型肝炎疫苗抗原工程菌的摇瓶发酵温度、初始pH值、溶解氧、IPTG浓度及诱导时间进行优化;以摇瓶发酵结果为基础,进一步对发酵罐培养工艺中诱导时机、诱导时间及补料培养基配方进行优化;以确定的罐发酵工艺条件连续发酵5批,验证该发酵工艺。结果最适摇瓶发酵工艺条件为:发酵温度为37℃,初始pH值为7.0,高溶氧,IPTG诱导浓度为0.05 mmol/L,诱导时间为4 h;最适罐发酵工艺条件为:菌体A600值约为40时开始诱导,诱导时间为4 h,补料培养基配方为葡萄糖25%,MgSO4.7H2O 0.5%。按照确定的工艺连续发酵5批,最终菌体A600均达50.0以上,目的蛋白的表达量均高于15%。结论优化的重组戊型肝炎疫苗抗原工程菌的发酵工艺稳定性良好,已达到中试生产规模的要求。Objective To optimize the fermentation process of recombinant E.coli for expression of hepatitis E（HE）vaccine antigen and lay a foundation of large-scale production of the vaccine.Methods The fermentation temperature,initial pH,dissolved oxygen,IPTG concentration and induction time were optimized using the expression level of target protein and final cell concentration as indicators for evaluation.The process in fermentor was developed based on the fermentation result in shake flask,in which the time point and duration for induction as well as the formula of supplement medium were further optimized.The optimized process was verified by production of five successive batches of HE vaccine.Results The optimal temperature,initial pH value,IPTG concentration and induction time for fermentation in shake flask were 37 ℃,7.0,0.05 mmol / L and 4 h respectively,and high dissolved oxygen concentration was selected.However,the optimal condition of fermentation process in fermentor was as follows： the recombinant E.coli was induced when the A600 value was about 40,and the time duration for induction was 4 h,while the medium A consisting of 25% glucose and 0.5% MgSO4·7H2O was served as supplement medium.Five batches of recombinant E.coli were fermented by the final process,of which all the A600 values reached more than 50,and the expression level of target protein was not less than 15%.Conclusion The optimized fermentation process showed high stability,which met the requirements for pilot production.

关 键 词：重组戊型肝炎疫苗 发酵 表达量 菌体浓度 

分 类 号：R373.2[医药卫生—病原生物学] R392.33[医药卫生—基础医学]