IRX1真核表达载体的构建及在胃癌细胞SGC-7901中的定位 |
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引用本文: | 刘卫仁,于颖彦,倪培华,吴建林,计骏,张佳年,陈雪华,刘炳亚,朱正纲. IRX1真核表达载体的构建及在胃癌细胞SGC-7901中的定位[J]. 中华实验外科杂志, 2005, 26(1): 884-886. DOI: 10.3760/cma.j.issn.1001-9030.2009.07.024 |
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作者姓名: | 刘卫仁 于颖彦 倪培华 吴建林 计骏 张佳年 陈雪华 刘炳亚 朱正纲 |
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作者单位: | 上海交通大学医学院附属瑞金医院消化外科研究所,200025; |
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基金项目: | 国家自然科学基金资助项目科技部"863"重大专项资助项目上海市科委重点基础研究项目上海市浦江人才计划项目 |
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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1443 bp编码序列,将PCR扩增产物连接人pGEM-Teasy载体,经测序确认序列无误后,亚克隆入pEGFP-N1载体,构建pEG-FP-IRX1真核表达载体.采用Lipofectamine 2000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.
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关 键 词: | 胃癌 真核表达载体 |
Construction of IRXI eukaryotic expression vector and its sub-cellular localization in SGC-7901 gastric cancer cell line |
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Abstract: | 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 in-serted 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-1RX1 fusion protein was clearly localized in nuclei of gastric cancer cells. Conclusion pEGFP-IRX1 expression vector was con-structed successfully, and the IRX1 protein was mainly observed in nuclei,which settled a basis for further functional study of gene IRX1 in gastric cancer. |
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Keywords: | IRX1Gastric carcinomaIRX1Eukaryotic expression vector |
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