重组人肝细胞生长因子重链在大肠杆菌中的表达研究  

Expression of recombinant human hepatocyte growth factor heavy chain in Escherichia coli

作　　者：党素英[1] 程牛亮[1] 牛勃[1] 王惠珍[1] 赵建滨[1] 

机构地区：[1]山西医科大学生物化学与分子生物学教研室,太原030001

出　　处：《山西医科大学学报》2000年第6期481-483,共3页Journal of Shanxi Medical University

摘　　要：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hHGFWT5”BZ] Objective To observe the expression of the human hepatocyte growth factor α chain cDNA,which was processed and modified before transcription,and then cloned into expression vector pBV220.To select the optimum conditions for expression of exogenous gene and to establish effective prokaryotic expression system for hHGF α protein. Methods and Results The expression plasmoid pBV220 hHGF α was transformed into E.coli DH 5α and Plyss.The culture temperature was changed from 30 ℃ to 42 ℃ to induce the expression of the goal protein.The expression of HGF α chain was tested by SDS PAGE.The result showed an obvious protein band,molecular weight in consistent with monobody HGF α chain ,appeared in the positive sample lane.Densitometric scaning of the SDS PAGE mapping was adopted to evaluate the amount of the goal protein.It was indicated that the product of the exogenous gene was 25% of total bacterial soluble protein of DH 5α hHGF α expression system and 30% of the plyss hHGF α expression system.Then the product was further identified by Western blot.A positive reaction of the product of the exogenous gene with the specific antibody of HGF α protein was observed. Conclusion The E.coli expression system,an effective source of providing HGF α protein in vitro ,is perfectly designed and completed. [WT5”HZ]

关 键 词：肝细胞生长因子 基因表达 大肠杆菌 hHGF重链 

分 类 号：Q786[生物学—分子生物学]