HMGB1基因真核细胞表达载体的构建及其在人脐静脉血管内皮细胞中的表达  

Construction of a Recombinant Eukaryotic Expression Plasmid Containing Human HMGB1 Gene and Its Expression in Human Umbilical Vein Endothelial Cells

作　　者：张晓娟[1] 李旭[1] 栾正刚[1] 马晓春[1] 

机构地区：[1]中国医科大学附属第一医院重症医学科,沈阳110001

出　　处：《中国医科大学学报》2012年第2期143-145,共3页Journal of China Medical University

基　　金：国家自然科学基金资助项目(30901438/C160402)

摘　　要：目的构建HMGB1基因真核细胞表达载体并检测其在真核细胞中的表达。方法提取人脐静脉血管内皮细胞(HUVEC)总RNA,反转录为cDNA。PCR扩增HMGB1全长编码基因,构建成pcDNA-3.1-myc-his-B-HMGB1真核细胞表达载体。转染HUVEC,Western blot检测转染后HMGB1基因的表达改变。结果构建HMGB1全长基因真核细胞表达载体成功,酶切鉴定及测序鉴定表明载体构建正确,转染HUVEC后可检测到外源性蛋白的表达,对照组无外源性蛋白表达。结论成功构建了HMGB1全长基因真核细胞表达载体并可转染至HUVEC中。Objective To construct a recombinant eukaryotic expression plasmid containing human HMGB1 gene and express the gene in human umbilical vein endothelial cells（HUVECs）.Methods Total RNA was extracted from HUVECs.The HMGB1 coding sequence was amplified by polymerase chain reaction（PCR） method and subcloned into pcDNA-3.1-myc-his-B vector.After the target region was sequenced,the plasmid was transfected into HUVECs.The expression of the recombinant plasmid in HUVECs was proved by Western blot.Results HMGB1 protein had been constructed into the pcDNA-3.1-myc-his-B expressing vector successfully.The fragment lengths were（647 bp and 5 500 bp） identified by double enzymes digestion.The expression of HMGB1 protein was detected by Western blot.Conclusion A recombinant eukaryotic expression plasmid containing HMGB1 gene was successfully constructed and expressed in HUVECs.

关 键 词：高迁移率族蛋白1基因 人脐静脉血管内皮细胞 真核细胞表达载体 转染 

分 类 号：R364.5[医药卫生—病理学]