论文部分内容阅读
目的 观察经过转录前加工、修饰后克隆至表达载体pBV2 2 0中的人肝细胞生长因子α链cDNA在大肠杆菌的表达情况 ,优化表达条件 ,以建立hHGFα原核高效表达体系 ,并进一步研究hHGF -α蛋白的生物学功能。方法与结果 将重组表达质粒pBV2 2 0 hHGF α转化大肠杆菌DH5α、Plyss ,通过升温诱导法 ,即将温度由 30℃升高到 42℃ ,诱导外源基因表达。SDS 聚丙烯酰胺凝胶电泳检测hHGF α蛋白表达 ,电泳图谱显示一特异的分子量与单体hHGF α链吻合的蛋白带。SDS PAGE光密度扫描对外源基因表达产物进行初步定量 ,表达产物分别占细菌可溶性蛋白总量的 2 5 %、30 %。进一步行West ern blot对表达产物进行定性 ,外源基因表达蛋白与特异的hHGF α抗体呈阳性反应。结论 成功地建立了hHGF α链原核高效表达工程菌系
Objective To observe the expression of human hepatocyte growth factor - α cDNA cloned into the expression vector pBV220 after pre - transcriptional modification, and to optimize the expression conditions in order to establish the prokaryotic expression system of hHGFα and to further investigate the expression of hHGF - Biological function of alpha protein. Methods and Results The recombinant plasmid pBV2 20 hHGF α was transformed into E. coli DH5α and Plyss. The temperature was raised from 30 ℃ to 42 ℃ and the expression of exogenous gene was induced by heat-induced induction. SDS polyacrylamide gel electrophoresis hHGF α protein expression, electrophoresis pattern showed a specific molecular weight and monomeric hHGF α chain anastomosis protein bands. SDS PAGE optical density scanning of foreign gene expression products were initially quantified, the expression product accounted for 25% of the total bacterial soluble protein, 30%. Western ern blot was further used to characterize the expression product, and the exogenous gene expression protein was positive with specific hHGF α antibody. Conclusion The prokaryotic expression system of prokaryotic hHGF alpha chain was established successfully