An infectious and selectable full-length replicon system with hepatitis C virus JFH-1 strain

Abstract

Aim: The hepatitis C virus (HCV) strain JFH-1 was cloned from a patient with fulminant hepatitis. A JFH-1 subgenomic replicon and full-length JFH-1 RNA efficiently replicate in cultured cells. In this study, an infectious, selectable HCV replicon containing full-length JFH-1 cDNA was constructed.

Methods: The full-genome replicon was constructed using the neomycin-resistant gene, EMCV IRES and wild-type JFH-1 cDNA. Huh7 cells were transfected with RNA synthesized in vitro, and then cultured with G418. Independent colonies were cloned to establish cell lines that replicate the full-length HCV replicon.

Results: HCV RNA replication was detected in each isolated cell line. HCV proteins and HCV RNA were secreted into culture medium, and exhibited identical density profiles. Interestingly, culture supernatants of the replicon cells were infectious for naïve Huh7 cells. Long-term culture did not affect replication of replicon RNA in the replicon cells, but it reduced core protein secretion and infectivity of culture supernatant. Culture supernatant obtained after serial passage of replicon virus was infectious for Huh7 cells.

Conclusions: Selectable infection was established using HCV replicon containing full-length genotype 2a JFH-1 cDNA. This system might be useful for HCV research.