摇瓶中重组大肠杆菌生产人γ-干扰素工艺优化  

A study on the fermentation technology of recombinant E.coli expressing human interferon-γ (hIFN-γ) in shaking flash

作　　者：申烨华[1] 刘江峰[1] 王骊丽[1] 耿信笃[1] 

机构地区：[1]西北大学现代分离科学研究所/现代分离科学陕西省重点实验室,陕西西安710069

出　　处：《西北大学学报（自然科学版）》2006年第4期571-574,共4页Journal of Northwest University（Natural Science Edition）

基　　金：国家自然科学基金资助项目(296701739880003)

摘　　要：目的优化大肠杆菌生产重组人γ-干扰素摇瓶发酵工艺条件，分析摇瓶发酵过程中的制约因素。方法在摇瓶中研究了种子菌龄、接种量和诱导时机等因素对工程菌生产rhlFN-γ和质粒稳定性的影响，及优化条件下工程茵发酵参数。结果最佳条件：种子液为OD6000．5—1．5，接种量为6％，培养至OD600为1．5时42℃诱导表达4h。在此条件下，在摇瓶中使用M9II培养基时，质粒无丢失，摇瓶中rhlFN-γ的表达量占菌体总蛋白的43．2％，光密度为3．3。对发酵过程中总糖、还原糖、总氮和pH的分析表明，发酵过程中培养基中碳源、氮源丰富，而溶解氧的不足和pH值偏酸性可能是整个培养过程的限制性因素。结论构建的工程菌发酵的稳定性和重复性良好，为rhlFN-γ的大规模生产提供可靠的放大依据。Abstract： Aim To optimize fermentation technology of recombinant E. Coli expressing human interferon-γ （hIFN-γ） with shaking flask. To analyze the unfavorable factors in the whole fermentation process. Methods The effects of inoculum age, inoculum volume, the time of increasing temperature to 42 ℃ and so on were investigated for the yield of rhIFN-γ and stability of plasmid. The characteristic parameters of fermentation by shaking flask were analyzed. Results The experimental results show that the optimum conditions are： inoculum age 0. 5 - 1.5 （ A600 ）, inoculum volume 6%. When cell density reached to OD600 up to 1.5, the temperature was increased up to 42℃ and continued for 4 h. Under these culture conditions, the plasmid didn＇t lose, and the expression and optical density （ A600） in shaking flake were 43.2% and 3.3, The results of characte ristic parameters of fermentation by respectively when the optimized MglI medium was used. shaking flask show that carbon and nitrogen sources were rich in MglI medium, but the dissolved oxygen was insufficient and the pH was acidic, which may be the unfavorable factors in the whole fermentation process. Conclusion The results indicate that the rhlFN-γ engineered E. Coli strain has better stability and repeatability which provides reliable data for mass production of rhlFN-γ.

关 键 词：重组人Γ-干扰素 重组大肠杆菌 摇瓶发酵 

分 类 号：TQ920.6[轻工技术与工程—发酵工程]