摘要
目的:研究丙型肝炎病毒(HCV)核心蛋白(core)反式激活基因HCTP4基因序列表达的调控机制。方法:选取翻译起始密码子ATG上游441 bp的DNA序列为启动子序列,应用聚合酶链反应技术(PCR),以肝母细胞瘤细胞系HepG2基因组DNA为模板,扩增该启动子DNA片段,将其克隆至pCAT3中,构建pCAT3-HCTP4-promoter报告基因表达载体,以该质粒转染HepG2细胞,用酶联免疫吸附方法(ELISA)检测报告基因编码产物氯霉素乙酰转移酶(CAT)的表达活性。结果:发现质粒pCAT3-HCTP4-promoter能够指导CAT的表达,吸光度(A值)是pCAT3对照质粒的5倍。结论:本研究克隆的启动子DNA序列具有转录活性,这一结果为研究HCTP4的调节机制,进一步阐明丙型肝炎病毒核心蛋白的作用机制奠定了基础。
Objective: To clarify the expression and regulation mechanism of the new target gene (HCTP4) transactivat-ed by core protein of hepatitis C virus (HCV) .Methods: Four hundred and forty-one base pairs (bp) upstream of the translation start site was selected as promoter sequence which was amplified from hepatoblastoma cell line-HepG2 cell genomic DNA by polymerase chain reaction (PCR) .The amplified product was cloned into pCAT3 vector. The HepG2 cell line was tranfect-ed by pCAT3-HCTP4-promoter.The CAT activity was detected by an enzyme-linked immunoassay (ELISA) kit.Results: We found pCAT3-HCTP4-promoter had activity of CAT. The expression of CAT was 5 times as higher as pCAT3-basic control vector. Conclusion: These results indicated that the pCAT3-HCTP4-promoter has promoter activity.
出处
《中西医结合肝病杂志》
CAS
2003年第6期354-356,共3页
Chinese Journal of Integrated Traditional and Western Medicine on Liver Diseases
基金
国家自然科学基金(No.C39970674
No.C03011402
No.C39900130
No.C30070689)
