精氨酸激酶基因的蛋白表达及实时荧光定量分析

Protein Expression of Arginine Kinase and Real-Time PCR Analysis

为构建棉铃虫精氨酸激酶基因(AK)的原核表达系统及确定2种不同品系的棉铃虫体内AK的表达量差异,以pET-28a质粒为表达载体,将AK基因重组转化至大肠杆菌BL21中进行原核表达。采用SDS-PAGE电泳及实时荧光PCR定量分析。结果表明:经IPTG诱导AK基因在大肠杆菌BL21得到表达,表达的融合蛋白其分子量与预测结果一致;石河子地区田间品系棉铃虫体内AK基因的表达量远远大于敏感品系棉铃虫体内AK基因的表达量为77.36%,表明石河子田间品系棉铃虫的生命代谢活动的能力要强于敏感品系。

In order to construct prokaryotic expression vector and determine AK expression quantity difference in two strains of cotton bollworm,AK gene of cotton bollworm was cloned into pET-28a vector and then transferred into E.coli BL21（DE3）.SDS-PAGE electrophoresis confirmed that AK protein was produced in E.coli BL21（DE3） by IPTG induction.The molecular weight of the fused protein was consistent with the predicted molecular weight.Real-time PCR result showed that the AK expression quantity of cotton bollworm in the field population of Shihezi was significantly higher than that from susceptible population,indicating that the metabolic activity of cotton bollworm in the field population of Shihezi was stronger than that from susceptible population.