犬细小病毒VP2基因原核表达条件优化与蛋白纯化

Optimization and Purification of Prokaryotic Expression of Canine parvovirus VP2 Gene

为了提高犬细小病毒VP2基因在大肠杆菌E.coli BL21(DE3)中的表达量,通过改变诱导温度、诱导时间及诱导剂浓度等条件对表达量产生影响,以SDS-PAGE电泳分析证明VP2基因蛋白表达的最佳条件,并通过Glutathione Sepharose 4B亲和层析柱法纯化目的蛋白。结果表明,重组质粒E.coliBL21(DE3)在35℃,1.0 mmol/mL IPTG诱导6 h时条件下蛋白表达量最高。SDS-PAGE电泳检测的纯化目的蛋白约为90 kDa,与预计大小一致。VP2基因融合蛋白的优化表达和纯化为研究犬细小病毒亚单位疫苗奠定了基础。

In order to improve the expression level of Canine parvovirus（CPV） VP2 gene in Escherichia coli,optimal conditions for expression level of GST-VP2 fusion protein were analyzed by SDS-PAGE at different concentration,temperature and time of employed inducer（IPTG）.The protein was purified by glutathione sepharose 4 B affinity column.The results showed that the fusion protein expression of recombinant plasmid Escherichia coli,BL21（DE3） was the optimal when 1.0mmol/mL IPTG were induced for 6 h at 35 ℃.Fusion protein purified by Glutathione Sepharose 4B affinity column was about 90 kDa which were consistent with the expected size by SDS-PAGE.The optimized expression and purification of VP2 gene fusion protein set the base for studying sub-unit vaccine of CPV.