重组慢病毒载体建立稳定表达绿色荧光蛋白宫颈癌细胞系  

Establishment cervical cancer cell line with GFP expressing using replication incompetent lentiviral vectors

作　　者：苏桂栋[1] 尹倩[1] 孙桂芹[1] 刘柯伶[1] 张燕[1] 钟梅[1] 

机构地区：[1]南方医科大学南方医院妇产科,广东广州510515

出　　处：《中国实用妇科与产科杂志》2010年第11期833-836,共4页Chinese Journal of Practical Gynecology and Obstetrics

基　　金：广东省自然科学基金(编号:07005147)

摘　　要：目的构建含绿色荧光蛋白(GFP)基因的重组慢病毒,导入宫颈癌细胞系,建立稳定表达GFP细胞系。方法 2007年9月至2010年6月将含GTP基因载体质粒pGC-FU、包装质粒pHelper1.0及包膜质粒pHelper2.0,用磷酸钙共沉淀法共转染包装细胞系293T,收集病毒颗粒感染人宫颈癌Siha细胞,以流式细胞仪筛选出荧光亮度较强GFP阳性的细胞继续培养。结果利用慢病毒载体可稳定转入GFP基因,获得稳定表达GFP的新宫颈癌细胞系S3,此细胞系中GFP阳性率达99%以上,比较Siha和S3两者生长曲线、细胞周期分布、HPV-16E7的表达量、裸鼠皮下成瘤能力差异均无统计学意义。结论慢病毒高效转入GFP基因,成功建立了表达GFP的S3细胞系,为宫颈癌基因研究提供了便利。Objectives To establish a green fluorescent protein （GFP） stably expressing human cervical cancer cell line using replication incompetent and to investigate the expression of GFP in the cells. Methods Vector plasmid pGC-FU containing GFP gene as a target gene , package construct pHelper 1.0 and an envelope plasmid pHelper 1.0 were co-transfected into 293 T packaging cell line using Ca-P co-precipitation： The viruses harvested from the supernatant were used to transfect cervical cancer cells. Then the cells that strongly expressed GFP was selected with FACS. Results A stable GFP-expressing call line $3 was established, in this cell line more than 99 % of the cells expressed GFP. There is no significant difference between Si- ha and ,53 inits growth curves, cell cycle distributions ,the quantity of expressing HPV-16E7 and tumorigenic abilities in nude mice. Conclusion These researches showed that the lentiviral vector was efficient to establish human GFP-expressing cervical cancer cell lines-S3 ,which could facilitate the research on gene regulation of cervical carcinoma.

关 键 词：宫颈癌 绿色荧光蛋白 慢病毒 

分 类 号：R71[医药卫生—妇产科学]