犊牛前胸腺素α基因在大肠埃希菌中的表达及活性分析

Expression of Calf Prothymosin-α(ProT-α) Genes in E. coli and Its Activities Analysis

以RT-PCR法扩增犊牛前胸腺素α基因(prothymosin-α,ProT-α),与原核表达载体pGEX-4T-1连接生成重组质粒pGEX/ProT-α,再将重组表达质粒转化大肠埃希菌BL21(DE3)。重组菌经IPTG诱导后表达的GST-ProT-α融合蛋白主要存在于细菌裂解液中。SDS-PAGE电泳表明,GST—ProT-α融合蛋白表达量较高,分子量为38 ku;Western-blot和动物细胞试验表明,该产物能与胸腺素α1抗体发生特异性免疫反应,并可显著提高小鼠脾细胞增殖率和NK细胞杀伤活性。

ProT-α genes of calf were clones by RT-PCR. Both the ProT-α genes and proearyotie expression vectors （ pGEX4T-1 ） were digested with EeoR I and BamH I, and linked each other to produee recombinant plasmid pGEX/ ProT-α, and converted E. coli BL21 （DE3） with it. After induced with IPTG （isopropyl-beta-D-thiogalaetopyrano- side） the fusion protein of GST-ProT-α expressed by the recombinant E. coil existed in the bacterial lytie solution. SDS-PAGE eleetrophoresis indicated that the fusion protein of GST-ProT-α was highly expressed in amount with molec- ular weight at 38 ku. Western-blot and animal experiment indicated that the product eould specifically reaet immunoreaction with ProT-α I antibody, and remarkably raise the proliferation of mouse spleen eell and NK eell killing activities.