小麦TaMBD2原核表达载体的构建和诱导表达

Construction of Prokaryotic Expression Vector for TaMBD2 and Its Induced Expression

构建了小麦甲基结合域蛋白基因TaMBD2的原核表达载体pGEX-TaMBD2,并在大肠杆菌BL21(DE3)工程菌株中优化了融合蛋白GST-TaMBD2的诱导表达条件。结果表明,用0.3、0.5和1.0mmol·L-1的IPTG诱导后,融合蛋白GST-TaMBD2均能有效表达,以1.0mmol·L-1 IPTG诱导的效果最好;从诱导表达的时间来看,3种浓度IPTG诱导1h后融合蛋白均开始表达,且表达量随着诱导时间的延长而逐渐增加,但在诱导6h后的表达量增加幅度不大,因此确定诱导融合蛋白GST-TaMBD2表达的最佳IPTG浓度为1.0mmol·L-1,诱导时间为6h。

A prokaryotic expression vector pGEX-TaMBD2 for the wheat MBD （methyl-binding domain protein） gene TaMBD2 was constructed and the induced expression conditions for the fusion protein GST-TaMBD2 were optimized in the engineered Escherichia coli BL21 （DE3）. The results showed that the fused GST-TaMBD2 protein could be effectively expressed under different concentrations of IPTG, including 0.3, 0.5 and 1.0 mmol·L^-1, and the most suitable concentration of IPTG was 1.0mmol·L^-1. As to the induction time, the fusion protein began to express after 1 h of induction under the 3 different IPTG concentrations, its expression abundance was obviously increased along with the induction time and reached to the highest point at the 6 h of induction; in general, the most suitable induction condition for the fusion protein was 6 h induction under 1.0 mmol·L^-1 IPTG.