丙型肝炎病毒核心蛋白与其结合蛋白HCBP12在HepG2细胞中的相互作用

Interaction between HCV core Protein and HCBP12 in HepG2 cells

目的:使用哺乳动物双杂交系统探讨HCV-core与HCBP12在人肝癌细胞HepG2细胞中的相互作用.方法:PCR分别扩增含HCV core及HCBP12的cDNA片段,克隆入pGEM-T载体,经测序正确后再分别克隆入哺乳动物双杂交的表达质粒PBIND和PACT中,并转染HepG2细胞,48h后检测细胞中报告基因luciferase activity的表达水平.结果:Dual-Luciferase Activety系统检测显示PBIND-core与PACT-HCBP12实验组萤火虫荧光与海肾素荧光的比值明显高于其他阴性对照组,但低于阳性对照组.结论:哺乳动物双杂交系统证实HCV核心蛋白与其结合蛋白HCBP12在人肝癌细胞系中HepG2中有一定的相互作用,为进一步研究HCV-core和HCBP12的功能奠定基础.

AIM： To analyze the interaction between HCV core protein and HCBP12 in HepG2 cells using MammalianTM Two-Hybrid System. METHODS： cDNA fragments encoding HCV core protein and HCBP12 were respectively amplified by PCR and subsequently cloned into pGEM-T vector. Mter verified by sequencing, they were subcloned into two hybrid plasmids, PBIND and PACT. Then, PBIND-core, PACT-HCBP12 and pGSluc, a reporter vector, all were co-transfected into HepG2 cells. After 48 hours, the interaction between HCV core protein and HCBP12 was assayed by measuring reporter gene expression. RESULTS： Dual-lucfferase Reporter Assay showed that the lucfferase activity of the co-transfecfion experiment group was higher than that of the other negative control groups and lower than that of the positive control group. CONCLUSION： Mammalian Two-Hybrid Assay confirmed that HCBP12 can bind with HCV core protein to some extent, which lays foundation for further study on function of HCBP12 and HCV-core.