鼠血管抑素Angiostating真核表达载体的构建及在人胃癌细胞中的表达  

Construction and expression of eukaryotic expression plasmid with mouse angiostatin cDNA in human gastric cancer cells

作　　者：吴静[1] 樊代明[1] 时永全[1] 苗继延[1] 

机构地区：[1]第四军医大学西京医院全军消化病研究所,陕西西安710033

出　　处：《第四军医大学学报》2001年第10期908-912,共5页Journal of the Fourth Military Medical University

摘　　要：目的　构建鼠源性血管抑素 angiostatin c DNA真核表达载体 ,转染人胃癌细胞株 SCG790 1,检测蛋白表达 .方法　采用定向克隆构建鼠源性血管抑素 angiostatin c DNA真核表达载体 pc DNA3.1(+) - angiostatin,酶切鉴定和测序 ;采用脂质体基因转染人胃癌细胞株 SCG790 1,G418抗性筛选 ;Western blot检测血管抑素的表达 .结果　酶切鉴定和测序证明成功构建了基因序列正确的血管抑素真核表达载体 ,经G418筛选和 Western blot检测得到高表达血管抑素的细胞株 .结论　真核表达载体 pc DNA3.1(+) - angiostatin转导的人胃癌细胞能够表达血管抑素蛋白 。AIM To construct angiostatin cDNA eukaryotic expression plasmid, transfect human gastric tumor cells SCG7901, and identify the expression of angiostatin. METHODS Angiostatin cDNA (plasminogen k1 k3 with signal peptides and HA tag) was subcloned into eukaryotic expression plasmid pcDNA3.1(+). Gene transfer into gastric tumor cells SGC7901 was carried out by liposome (Tfx TM ), according to the transfection protocol in the manual. Thenthrough G418 screening, stable resistant clones were obtained. Western blot was used to confirm the expression of angiostatin in selected clones. RESULTS The constructed plasmid pcDNA3.1(+) angiostatin were confirmed with the right gene sequence by enzymatic digestion and gene sequencing. Western blot showed that angiostatin protein was expressed and secreted by SGC7901 cells. CONCLUSION Gene of angiostatin in plasmid ,pcDNA3.1(+) angiostatin was expressed by transfected cells, which had potential value in angiogenesis gene theraopy of gastric cancer.

关 键 词：血管抑素 真核表达 脂质体转染 肿瘤血管生成 胃癌 Angiostating 

分 类 号：R735.2[医药卫生—肿瘤]