Transfection of a differentiated human hepatoma cell line (Huh7) with in vitro-transcribed hepatitis C virus (HCV) RNA and establishment of a long-term culture persistently infected with HCV.
T7 RNA polymerase transcripts of a putative full-length cDNA clone of hepatitis C virus type 1 (HCV-1) were used to transfect a differentiated human hepatoma cell line, Huh7. The transfected genome replicated in cells, as evidenced by the appearance of progeny HCV RNA, detection of negative-strand viral RNA, and incorporation of [3H]uridine into the viral genome. Incubation of naive Huh7 cells with conditioned medium from transfected cells resulted in a new HCV infection, suggesting the production of biologically active virus in the inoculum. Maintenance of the transfected cells under serum-free culture conditions resulted in the selection of persistently infected cells which displayed a distinctive cellular morphology. This is the first demonstration that HCV RNA produced from cloned HCV cDNA is infectious and replication competent. This approach should provide a valuable system for studying HCV replication, persistence, and pathogenicity.
Yoo
B J
BJ
Chiron Corporation, Emeryville, California 94608.
Selby
M J
MJ
Choe
J
J
Suh
B S
BS
Choi
S H
SH
Joh
J S
JS
Nuovo
G J
GJ
Lee
H S
HS
Houghton
M
M
Han
J H
JH
D001483:Base Sequence; D006528:Carcinoma, Hepatocellular; D016895:Culture Media, Serum-Free; D016174:Hepacivirus / Q000235:genetics* / Q000472:pathogenicity; D006801:Humans; D008969:Molecular Sequence Data; D012367:RNA, Viral / Q000235:genetics*; D014158:Transcription, Genetic; D014162:Transfection; D014407:Tumor Cells, Cultured
D016895:Culture Media, Serum-Free; D012367:RNA, Viral