摘要
目的 建立容易检测的丙型肝炎病毒 (HCV)细胞模型。方法 利用基因重组技术 ,构建HCVcDNA与荧光素酶 (luc)融合基因可调控逆转录病毒载体 ;通过脂质体介导方法转染人肝癌细胞 (HHCC) ,观察luc基因的表达。结果 ①成功地将luc基因融合于HCVC区和大部分E1区基因的下游 ,构建了带HCV luc融合基因的真核表达载体 (pBPST HCV luc) ;②该载体在细胞内有效表达luc活性 ,puromycin筛选可提高luc的表达水平 ,并可受四环素的调节。结论 初步建立了表达HCVC E1和luc融合基因的可调控转染细胞模型 ,为以HCVC
Objective To establish HCV cell culture model which is easy to measure. Methods Controllable retroviral vector with fusion gene of hepatitis C virus (HCV) cDNA and luciferase (luc) reporter gene was constructed by molecular cloning technique, the transfection of this retroviral vector in a human hepatic carcinoma cell (HHCC) line was performed by lipofectAMINE and then luciferase activity in the cellular lysate was measured by scintillation counter. Results ① Fusion gene of the HCV 5′ NCR C region and luciferase reporter gene identified by restriction endonuclease cleavage have been cloned into pBPSTR1 vector. ② The luciferase activity could maintain up to 20 days at least, and could be increased by puromycin treatment and regulated by tetracycline. Conclusion A cell model for expression of HCV C E1 and luciferase genes was established for gene therapy studies against HCV C E1 sequences.
出处
《中华微生物学和免疫学杂志》
CSCD
北大核心
2000年第2期167-169,共3页
Chinese Journal of Microbiology and Immunology
