IRX1真核表达载体的构建及在胃癌细胞SGC-7901中的定位  

Construction of IRX1 eukaryotic expression vector and its sub-cellular localization in SGC-7901 gastric cancer cell line

作　　者：刘卫仁[1] 于颖彦[1] 倪培华[1] 吴建林[1] 计骏[1] 张佳年[1] 陈雪华[1] 刘炳亚[1] 朱正纲[1] 

机构地区：[1]上海交通大学医学院附属瑞金医院消化外科研究所,200025

出　　处：《中华实验外科杂志》2009年第7期884-886,共3页Chinese Journal of Experimental Surgery

基　　金：国家自然科学基金资助项目（30572127;30770961;30670939）;科技部“863”重大专项资助项目（2006AA02A402;2006AA02A301）;上海市科委重点基础研究项目（05JC14013）;上海市浦江人才计划项目（PJ200700367）

摘　　要：目的构建pEGFP—IRX1真核表达载体，体外转导至SGC-7901胃癌细胞株，观察IRX1基因转染后蛋白表达与细胞内定位。方法PCR扩增IRX1全长1443bp编码序列，将PCR扩增产物连接入pGEM-Teasy载体，经测序确认序列无误后，亚克隆入pEGFP—N1载体，构建pEGFP-IRX1真核表达载体。采用Lipofectamine2000转染SGC-7901胃癌细胞株，在荧光显微镜下观察外源性IRX1基因导入后在胃癌细胞内的表达与亚细胞定位。结果成功构建有绿色荧光蛋白融合的IRX1真核表达载体，外源性IRX1基因在胃癌SGC-7901细胞中表达成功，在荧光显微镜下IRX1表达蛋白定位于细胞核内。结论pEGFP—IRX1真核表达载体构建成功，并可在细胞内表达，这将为IRX1在胃癌内的功能研究奠定基础。Objective To construct the eukaryotic expression vector of pEGFP-IRX1 ,and observe the IRX1 protein expression and its sub-cellular localization in SGC-7901 gastric cancer cell line. Methods The coding sequence of IRX1 1443 bp was obtained by PCR amplification,then the PCR product was inserted into the pGEM-Teasy vector. After confirming the DNA sequence, IRX1 amplification product was sub-cloned into pEGFP-N1 vector. Finally, an IRX1 gene eukaryotic expression vector Was obtained. The pEGFP-IRX1 expression vector was transfected into SGC-7901 gastric cancer cell line by Lipofectamine 2000,and the expression of pEGFP-IRX1 was detected by GFP fluorescence. Results The recombinant vector was highly expressed in SGC-7901 cells. The green fluorescence of EGFP-IRX1 fusion protein was clearly localized in nuclei of gastric cancer cells. Conclusion pEGFP-IRX1 expression vector was constructed successfully,and the IRX1 protein was mainly observed in nuclei,which settled a basis for further functional study of gene IRX1 in gastric cancer.

关 键 词：胃癌 IRX1 真核表达载体 

分 类 号：R735.2[医药卫生—肿瘤] R780.2[医药卫生—临床医学]