hLMO3基因全长及截短序列原核表达载体的构建及蛋白诱导表达  

Construction of prokaryotic plasmid of human LMO3 and deletions gene and identification of their recombinant proteins

作　　者：刘彤[1] 顾卉[2] 李洋[1] 李丰[1] 

机构地区：[1]中国医科大学基础医学院细胞生物教研室教育部医学细胞生物学重点实验室,沈阳110001 [2]中国医科大学附属盛京医院中心实验室,沈阳110001

出　　处：《解剖科学进展》2012年第1期8-11,共4页Progress of Anatomical Sciences

基　　金：国家自然科学基金重大研究计划(No.90813038)

摘　　要：目的构建hLMO3原核全长及5个截短表达载体并鉴定融合蛋白的表达。方法提取工具细胞HEK293的mRNA,反转录为cDNA,PCR扩增hLMO3全长及截短编码基因,亚克隆至GST融合表达载体pGEX-5X-2中。在E.coli BL21中诱导GST-hLMO3全长及截短融合蛋白表达,并经免疫印迹鉴定结果。结果 hLMO3全长及5个截短基因序列克隆到了原核表达载体pGEX-5X-2中,并经测序鉴定正确。Westernblot在E.coli Bl21中检测到了相应融合蛋白的表达。结论成功构建hLMO3全长及5个截短基因序列原核表达载体,在E.coli BL21中诱导表达出GST-LMO3全长及截短融合蛋白。Objective To construct an prokaryotic expression vector of LMO3（LIM-domain only 3） full-length and deletion gene and identify their recombinant protein expressions induced by isopro-pyl-1-thio-b-Dgalactopyranoside（IPTG）.Methods Total RNA was extracted from HEK293,hLMO3 and deletion coding sequences were amplified by polymerase chain reaction（PCR）method and cloned into pGEX-5X-2.The expression of GST-hLMO3 fusion proteins was induced by IPTG and identified by Western blot.Results The coding sequence of hLMO3 full-length and deletion gene were cloned into the pGEX-5X-2 which was transformed into BL21 successfuly,and identified by double enzymes digestion and sequencing.The expression of GST-hLMO3 fusion proteins induced by IPTG was observed in Bl21.Conclusion The recombinant prokaryotic plasmids were constructed successfully into pGEX-5X-2 and the expression of GST-LMO3 full-length and deletion fusion proteins was identified.

关 键 词：hLMO3 工具细胞HEK293 原核表达 截短蛋白 

分 类 号：Q291[生物学—细胞生物学]