摘要
以脂质体基因转移技术，将含ＨＳＶ１ＴＫ基因的重组逆转录病毒载体ｐＬＴＲＮＬ转染人肝癌细胞系ｈＨＣＣ。经抗生素Ｇ４１８选择，筛选出ＨＳＶ１ＴＫ基因转染的ｈＨＣＣ，再进行克隆化和亚克隆化后扩大培养。应用ＰＣＲ，ＲＴＰＣＲ及ｓｌｏｔｂｌｏｔ狭槽印迹等方法，对转染细胞基因组ＤＮＡ和总ＲＮＡ进行了鉴定。结果表明，ＨＳＶ１ＴＫ基因已整合入ｈＨＣＣ细胞基因组中，且有ＴＫ基因的ｍＲＮＡ转录。ＨＳＶ１ＴＫ基因稳定转染的ｈＨＣＣ的建立，为在体外和动物模型上对肝癌进行基因治疗奠定了实验基础。
The recombinant retroviral vector pLTRNL containing the HSV 1 TK gene was transfected into human hepatocellular carcinoma cell line hHCC by lipofectamine method. The transfected cells were selected with 800mg/L of geneticin(G418) for three weeks, and then cloned and subcloned. G418 resistant transfected cells were named hHCC/TK cells. The total RNA and genomic DNA of hHCC/TK or hHCC were extracted. These genomic DNA containing TK gene were identified by PCR. The total RNA containing mRNA of HSV 1 TK gene were identified by RT PCR and RNA slot blot methods. The results showed that the HSV 1 TK gene was integrated into the genomc of transfected cells and the mRNA of TK gene has been transcribed. The establishment of stable long term transfected human hepatocellular carcinoma cell line bearing HSV 1 TK gene could provide the possibility of experimental gene therapy for human hepatocellular carcinoma in vitro and in vivo .
出处
《细胞与分子免疫学杂志》
CAS
CSCD
1998年第1期26-28,共3页
Chinese Journal of Cellular and Molecular Immunology
关键词
肝肿瘤
基因治疗
胸苷激酶
HSV-1
pLTRNL
herpes simplex virus thymidine kinase retroviral vector hepatocellular carcinoma gene therapy
