缬氨酸转氨酶工程菌的构建及表达条件的优化

目的]构建缬氨酸转氨酶基因(avtA)的大肠杆菌工程菌,并优化其表达条件。方法]将avtA基因插入到载体pET32a中,构建表达质粒pET32a-avtA,通过纸层析检测酶活性,SDS-PAGE凝胶电泳检测目的蛋白,并通过考察培养基中蛋白胨浓度、IPTG诱导浓度和诱导时间来优化其表达条件。结果]成功构建了缬氨酸转氨酶基因的大肠杆菌工程菌,其最佳表达条件为:培养基中蛋白胨浓度为12g/L,IPTG浓度为0.4 mmol/L,诱导时间为8 h。结论]缬氨酸转氨酶工程菌具有良好的应用前景。

Construction and Optimization of Engineering Bacteria to Express Valine Aminotransferase

Objective] To construct E.coli engineering bacteria expressing valine aminotransferase and optimize the expression conditions.Method] The avtA gene was inserted into expression vector pET32a and the expression plasmid pET32a-avtA was constructed.Valine aminotransferase of enzyme activity was detected by paper chromatography.The expression product of avtA gene was identified by SDS-PAGE,and the expression condition was optimized through inspecting the peptone concentration,IPTG concentration and induction time.Result] An E.coli engineering bacteria to express valine aminotransferase was successfully established.The optimum expression conditions were established as follows : 12 g/L of peptone,0.4 mmol/L of IPTG and 8 h induction time.Conclusion] Valine transaminase engineered bacteria had a good prospect.