rHAP在大肠杆菌中的诱导条件优化

Optimization of inducing conditions on rHAP producing E.coli

目的研究用乳糖替代异丙基-β-D-硫代半乳糖苷(IPTG)作为诱导剂进行重组rHAP的表达。方法研究了rHAP基因工程大肠杆菌的生长、工程菌表达rHAP时IPTG诱导表达的起始工程菌浓度、诱导物IPTG和乳糖的用量及乳糖诱导时间与rHAP表达水平之间的关系。结果在摇瓶实验中,通过优化,选择在培养物的A560值达到3.0时进行诱导剂诱导可以使重组蛋白的表达实现最大化;使用乳糖进行诱导,在浓度0.5 g/L时可以使rHAP的表达量与1 mmol/L的IPTG诱导时相当;诱导时间控制在3 h可使目的蛋白的产生量达到最大,能够占到总可溶性蛋白的20%。结论获得了使用乳糖作为诱导剂获得rHAP的最适诱导条件,降低了成本,为规模化生产打下了基础。

Purpose To study the inducing factors influencing the expression of rHAP by recombinant Escherichia coli strain, using lactose as an inducer instead of IPTG. Methods The relationship between bacterial growth and density,the dosage of inducer IPTG and lactose,the induction time and the expression level of recombinant protein were investigated. Results The optimized inducing time was found to be that when the concentration of the culture reached A560 value 3.0. Induced by lactose at a concentration of 0.5 g/L the expression of rHAP was the same as that induced by IPTG. The time of the induction could affect the expression level of the target protein. Conclusion The optimized inducing conditions were obtained to express rHAP in E. coli. In the manufacture of such a recombinant protein, using lactose instead of IPTG can reduce the cost. It could lay a foundation for production on larger scale.