Flavivirus-based system for production of hepatitis c virus (hcv)

Abstract

Provided herein is a mammalian cell transformed to contain a plasmid encoding a T7 or SP6 promoter operably linked to one or more HCV genes, a subgenomic replicon from a flavivirus and a cytoplasmic T7 and SP6 RNA amplification system. Also provided herein are isolated replication-competent HCV particles produced by the method comprising the steps of providing a transformed mammalian cell according to the first embodiment, culturing the cell, and recovering the replication-competent HCV particles from the cell culture. Provided herein are isolated HCV structural proteins produced by the method comprising the steps of providing a transformed mammalian cell according to the first embodiment, culturing the cell, and recovering the HCV structural proteins from the cell culture. Further provided herein is a system for assaying HCV entry into a cell comprising a first plasmid encoding a T7 or SP6 promoter operably linked to an HCV polynucleotide comprising at least the 5'-UTR to NS2 operably linked to an EMCV IRES in frame with an SP6 or T7 polymerase gene, respectively, a first host cell line expressing a replicon from a flavivirus and comprising a cytoplasmic T7 and SP6 RNA amplification system, a second plasmid encoding a reporter gene operably linked to both T7 and SP6 promoters in tandem, and a second host cell line comprising a cytoplasmic T7 polymerase or SP6 polymerase RNA amplification system.