重组人抗HBsAg单链抗体工程菌的中试发酵工艺研究

Study on the fermentation condition of recombinant Escherichia coli producing human anti-HBsAg scFv

目的研究表达重组人抗HBsAg单链抗体工程菌的中试发酵工艺。方法首先采用摇瓶培养,对培养基配方、pH值、诱导表达时机和诱导剂量进行优化,确定基本发酵参数,然后在30L发酵罐上进行中试规模发酵培养。结果在摇瓶培养时,发现含有甘油和微量元素等成分的改良培养基效果明显优于LB培养基和2×YT培养基,另外,工程茵的最佳pH值为7.0,最佳诱导时机为对数中期,最佳诱导剂浓度为0.4mmoL/L异丙基-1-硫代-β-呋喃半乳糖,最佳诱导时间为4h,将摇瓶培养确定的关键参数应用到30L发酵罐中,控制溶氧大于30%,进行3批发酵试验,发现发酵产物的湿菌产量可达58-61.9g/L,目的蛋白质表达量可达28.9%-31.2%以上。结论建立了工程菌M15[pQE-scFv]的中试发酵工艺,为进一步开发打下良好基础。

Purpose To develop a fermentation procedure on Escherichia coli M15 expressing anti-HBsAg scFv.Methods The effects of the composition of the medium, the fed-batch, the range of pH,induction time and induction dose were analyzed with shake flash. And the defined fermentation parameter was applied to 30 liter fermentation tank in a pilot scale.Results It was found that culture medium C with glycerol and microelement was better than LB and 2×YT,and optimum pH 7.0,optimum induction time log midphase,optimum induction dose 0.4 mmol/L IPTG,and optimun induction time 4 h.Under the established conditions,the yield of bacteria block reached 58-61 .9 g/L,and expressing level of recombinant anti-HBsAg scFv was about 28 .9% -31.2% of the host protein in 30 liter fermentation tank . Conclusion The fermentation process of engineering bacterium M15[pQE-scFv] was established.