title: A recombinant replication-competent hepatitis C virus expressing Azami-Green, a bright green-emitting fluorescent protein, suitable for visualization of infected cells.
abstract: The hepatitis C virus (HCV) production system consists of transfecting the human hepatoma cell line Huh7 with genomic HCV RNA (JFH1). To monitor HCV replication by fluorescence microscopy, we constructed a recombinant HCV clone expressing Azami-Green (mAG), a bright green fluorescent protein, by inserting the mAG gene into the nonstructural protein 5A (NS5A) gene; the resultant clone was designated JFH1-hmAG. The Huh-7.5.1 (a subclone of Huh7) cells transfected with JFH1-hmAG RNA were found to produce cytoplasmic NS5A-mAG, as readily visualized by fluorescence microscopy, and infectious virus, as assayed with the culture supernatant, indicating that JFH1-hmAG is infectious and replication-competent. Furthermore, the replication of this virus was inhibited by interferon alpha in a dose-dependent manner. These results suggest that JFH1-hmAG is useful for studying HCV life cycle and the mechanism of interferon's anti-HCV action and for screening and testing new anti-HCV drugs.
authors.affiliation: China-Japan Joint Laboratory of Molecular Immunology and Molecular Microbiology, Institute of Microbiology, Chinese Academy of Sciences, Chaoyang District, Beijing, China.
authors.forename: Wei
authors.initials: W
authors.lastname: Hou
authors.forename: Chie
authors.initials: C
authors.lastname: Aoki
authors.forename: Lijuan
authors.initials: L
authors.lastname: Yu
authors.forename: Xianzi
authors.initials: X
authors.lastname: Wen
authors.forename: Yinhuan
authors.initials: Y
authors.lastname: Xue
authors.forename: Bin
authors.initials: B
authors.lastname: Gao
authors.forename: Wenjun
authors.initials: W
authors.lastname: Liu
authors.forename: George Fu
authors.initials: GF
authors.lastname: Gao
authors.forename: Aikichi
authors.initials: A
authors.lastname: Iwamoto
authors.forename: Yoshihiro
authors.initials: Y
authors.lastname: Kitamura
mesh_terms: D000595:Amino Acid Sequence; D002460:Cell Line; D049452:Green Fluorescent Proteins / Q000032:analysis* / Q000235:genetics; D016174:Hepacivirus / Q000187:drug effects / Q000235:genetics / Q000502:physiology*; D006801:Humans; D016898:Interferon-alpha / Q000494:pharmacology; D008856:Microscopy, Fluorescence / Q000379:methods*; D014162:Transfection; D014779:Virus Replication / Q000187:drug effects
chemical_list: D016898:Interferon-alpha; D049452:Green Fluorescent Proteins