Molecular characterization and fluorescence analysis of HCV non-structural proteins NS3, NS3-4A and NS4A of genotype 3a.

Hepatitis C virus (HCV) chronically infects almost 2% of world's population. Chronic infection can lead to liver failure and hepatocellular carcinoma (HCC). Approximately 10% of the Pakistani population is infected with HCV and type 3 is the most prevalent genotype with 75-90% prevalence. In this study we have developed transiently expressing cell culture based system for the expression of HCV non-structural NS3, NS3-4A and NS4A proteins of genotype 3a. HCV non-structural genes NS3, NS3-4A and NS4A were cloned in to pFLAG-CMV2 and pEGFP-C1vectors. All vectors were transfected separately to Huh-7 cells and their protein expression was analyzed by Western blot and immunofluorescence. All proteins were expressed correctly and in the transfection we have obtained 42-70% efficiency for all clones. This system can be used for the development of novel antiviral strategies to inhibit the viral replication, to study apoptosis pathways induced by HCV, for the evaluation of vaccine candidates and also to study the role of HCV different signaling pathways.