Construction Of Plasmid Including HCV NS3 Gene And Expression Of NS3 Protein In Eukaryotic Cells

Aim: To construct eukaryotic plasmid which includes HCV NS3 gene, andinduce the expression of NS3 protein in human hepatocytes.Methods: HCV NS3 gene was extended by using PCR from plasmidpBRTM/HCV1-3011 which includes the full length of HCV gene. Werecombined the NS3 segment with expression plasmid pcDNA3.1( —) toconstruct eukaryotic pcDNA3.1( — )/NS3 plasmid. Then plasmidpcDNA3.1(—)/NS3 was transfected into human hepatocytes QSG7701 byusing poly-cation. And the expression of NS3 protem was detected byimmunohistochemical staining method. The cytoplasm protein was extractedfrom cells screened by G418, which can express NS3 protein steadily, and hadbeen analysed by SDS—PAGE and Western Blotting.Results: The cloned NS3 segment had correct length and sequence . TheQSG7701 cells were successfully transfected and expressed NS3 proteinwith right molecular weight and antigenic speciality.Conclusion: We have successfully constructed eukaryotic expression plasmidpcDNA3.1(-)/NS3 which includes the HCV NS3 gene. And the vectorsuccessfully expressed NS3 protein in human hepatocytes. These results getpreparation for further research in the mechanism of cancerization inducedby HCV NS3 gene in immortalized human hepatocytes.