摘要
目的探讨含anti-HBsAg Fab/pBAD表达载体的工程化大肠杆菌的大规模培养条件。方法在摇瓶发酵条件下探讨工程菌的生长和表达规律,摸索最佳的培养模式及诱导表达条件。后根据摇瓶发酵结果于发酵罐中行补料高密度发酵试验,确定最佳补料模式。结果由摇瓶发酵获得的数据表明,以培养体系处于对数生长中期为诱导表达的起点、在25 ℃条件下以0.2% 阿拉伯糖诱导12 h,Fab的得率最佳,采用溶氧控制补料模式可使培养体系的最大D600达到55.2,相当于湿菌含量110 g/L水平;同时,经证实Fab的抗原结合活性良好。结论初步确定了周期短、产率高且稳定可靠的发酵工艺路线,为应用原核表达体系工业化大批量生产基因工程抗体奠定了基础。
Objective To define the conditions for large-scale production of genetically engineered E.coli bearing humanized anti-HBsAg Fab. Method Characteristic growth and expression of the engineered E.coli were observed during fermentation in the shaking flask to define the optimal culture conditions to achieve the highest production levels. On the basis of the observation results, the E.coli was cultured in a fermentor using the fed-batch method to determine the optimal production techniques. Results Observation of the bacterium in the shaking flask showed initiation of the induction procedure in the mid-log growth phase at 25 ℃ with 0.2% arabinose resulted in the highest production of anti-HBsAg Fab. The D600 value of the culture reached 55.2, equivalent to 110 g/L wet weight of the bacterium, using the DO-stat fed-batch method. The resultant Fab showed well-preserved biological activity. Conclusion Reliable techniques for rapid and massive production of the Fab have been developed.
出处
《第一军医大学学报》
CSCD
北大核心
2004年第5期517-520,共4页
Journal of First Military Medical University
基金
国家自然科学基金(39670668)~~
