A Novel Hepatitis C Virus NS5B Polymerase Assay of De Novo Initiated RNA Synthesis Directed from a Heteropolymeric RNA Template

The hepatitis C virus (HCV) NS5B RNA-dependent RNA polymerase is essential for viral replication and a clinically validated antiviral target. Discovery of HCV polymerase inhibitors is greatly facilitated by the availability of a suitable biochemical assay using purified NS5B. We describe here a novel NS5B polymerase assay of de novo initiated RNA synthesis directed from a synthetic heteropolymeric RNA template modified with dideoxycytidine at its 3′-end. This assay has been successfully used for screening and characterization of both initiation and elongation inhibitors of the HCV NS5B polymerase.