Cell culture and infection system for hepatitis C virus.
Hepatitis C virus (HCV) infection causes chronic liver disease and is a worldwide health problem. Despite ever-increasing demand for knowledge on viral replication and pathogenesis, detailed analysis has been hampered by a lack of efficient viral culture systems. We isolated HCV genotype 2a strain JFH-1 from a patient with fulminant hepatitis. This strain replicates efficiently in Huh7 cells. Efficient replication and secretion of recombinant viral particles can be obtained in cell culture by transfection of in vitro-transcribed full-length JFH-1 RNA into Huh7 cells. JFH-1 virus generated in cell culture is infectious for both naive Huh7 cells and chimpanzees. The efficiency of viral production and infectivity of generated virus is substantially improved with permissive cell lines. This protocol describes how to use this system, which provides a powerful tool for studying viral life cycle and for the construction of antiviral strategies and the development of effective vaccines. Viral particles can be obtained in 12 days with this protocol.
Kato
Takanobu
T
Liver Diseases Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland 20892, USA.
Date
Tomoko
T
Murayama
Asako
A
Morikawa
Kenichi
K
Akazawa
Daisuke
D
Wakita
Takaji
T
D000328:Adult; D000818:Animals; D018929:Cell Culture Techniques / Q000379:methods*; D002460:Cell Line; D018274:Electroporation; D018014:Gene Transfer Techniques; D015386:Hazardous Substances; D016174:Hepacivirus / Q000302:isolation & purification / Q000502:physiology*; D006526:Hepatitis C / Q000821:virology; D006801:Humans; D008297:Male; D002679:Pan troglodytes; D012367:RNA, Viral; D014779:Virus Replication / Q000502:physiology*
D015386:Hazardous Substances; D012367:RNA, Viral