高密度培养大肠杆菌生产重组钩端螺旋体DNA疫苗

High cell-density culture of recombinant E.coli to produce leptospira DNA vaccine pcD-flaB

目的 :高密度、高质粒拷贝数培养大肠杆菌生产重组 DNA疫苗 pc D- fla B。 方法 :应用 FU S- 5 L自控发酵罐 ,采用补料分批培养技术 ,控制补料的补加使溶氧控制在 30 %～ 6 0 % ,p H控制在 7左右 ,培养中后期采用 4 2℃培养以提高质粒拷贝数。 结果 :重组菌 E.coli DH5α/ pc DNA 3- fla B发酵液光密度 (D6 0 0 )达到 4 5。经纯化后最终质粒 DNA得率为 5 0 m g/ L。质粒没有发生丢失现象。 结论 :本实验为钩端螺旋体 DNA疫苗的大规模生产奠定了基础。

Objective:To culture recombinant E.coli in high cell density with high plasmid copy number to produce DNA vaccine pcD flaB.Methods:FUS 5L type autocontrol fermentor and fed bath culture technology were used in this experiment, dissolved oxygen was kept at 30% 60% by fed batch control and pH was controlled at about 7 throughout the culture, 42℃ culture was used to raise copy number of plasmid at mid and later period to produce recombinant E.coli DH5α/pcDNA3 flaB. Results:The final cell density was 45.The yield of purified plasmid DNA was 50 mg/L. No plasmid was lost.Conclusion:This study provides foundation for large scale production of leptospira DNA vaccine.