Abstract: Objective:To investigate the transactivating effect of hepatitis C virus （HCV) nonstructural protein 5A （NS5A).Methods: HCV NS5A gene was amplified from plasmid pBRTM3011 and the amplified product was cloned into pcDNA3.1(-) vector. Then the hepatoblastoma cell line HepG2 was transfected by pcDNA3.1(-)-NS5A, and pcDNA3.1(-)-NS5A and reporter plasmid pCAT3 promoter, respectively.HCV NS5A protein expressed in HepG2 cells was detected by Western blotting method. The activity of CAT was detected by a ELISA kit, which reflect the transactivating function of HCV NS5A protein. Results: HepG2 cells transfected with pcDNA3.1(-)-NS5A can express HCV NS5A protein. The expression of CAT in HepG2 cells transfected with the pcDNA3.1(-)-NS5A was 3.8 times as higher as that of control plasmid. Conclusion: It is suggested that the recombinant plasmid pcDNA3.1(-)-NS5A can be expressed in mammalian cell line, and has transactivating effect on SV40 early promoter.