pGEX-5X-1-hLMO4原核质粒构建及重组蛋白表达  

Construction of pGEX-5X-1-hLMO4 prokaryotic plasmid and identification of its recombinant protein

作　　者：张红艳[1] 李彦姝[1] 姚远[2] 王春玉[1] 王迪[1] 李丰[1] 

机构地区：[1]中国医科大学细胞生物学教研室卫生部细胞生物学重点实验室教育部医学细胞生物学重点实验室,辽宁沈阳110001 [2]辽宁省人民医院消化内科,辽宁沈阳110016

出　　处：《东南大学学报（医学版）》2012年第2期139-143,共5页Journal of Southeast University(Medical Science Edition)

基　　金：国家自然科学基金资助项目(31171360;30800415);博士点基金资助项目(20102104110016)

摘　　要：目的:构建hLMO4的原核表达载体并诱导、纯化和鉴定其表达。方法:hLMO4全长编码基因经BamHⅠ和XhoⅠ双酶切后,克隆至GST融合表达载体pGEX-5X-1,异丙基β-D硫代半乳糖苷(IPTG)在BL21大肠杆菌中诱导GST-hLMO4融合蛋白表达,利用Glutathione Sepharose 4B纯化诱导的融合蛋白,并经Western blot鉴定结果。结果:hLMO4编码序列克隆至pGEX-5X-1载体中,双酶切鉴定片段大小为500 bp,在E.coli BL21中IPTG诱导融合蛋白的表达,分子质量约为49 000 Da,成功纯化出GST及GST-hLMO4蛋白,Western blot检测到蛋白表达。结论:构建了hLMO4基因原核表达载体,鉴定了GST-hLMO4融合蛋白表达。Objective：To construct prokaryotic expression vector of human LMO4 gene and induce,purify and identify its recombinant protein expression.Methods：The hLMO4 coding sequence was digested with BamHⅠ and XhoⅠ enzymes,and cloned into pGEX-5X-1.The expression of GST-hLMO4 fusion protein was induced by IPTG and identified by Western blot.Results：The coding sequence of hLMO4 gene was cloned into the pGEX-5X-1 plasmid which was transformed into E.coli BL21.The length of fragment was 500 bp.The expression of GST-hLMO4 fusion protein was induced by IPTG,and the molecular weight of protein was 49 000 Da.Conclusion：The recombinant prokaryotic plasmid was successfully constructed into pGEX-5X-1.The expression of GST-LMO4 fusion protein was induced by IPTG and identified.

关 键 词：LMO4 蛋白纯化 融合蛋白 

分 类 号：Q291[生物学—细胞生物学] Q591.2