Cell culture and infection system for hepatitis C virus

被引：161

作者：

Kato, Takanobu

Date, Tomoko

Murayama, Asako

Morikawa, Kenichi

Akazawa, Daisuke

Wakita, Takaji ^{[1
]}

机构：

[1] NIH, NIDDK, Liver Dis Branch, Bethesda, MD 20892 USA

[2] Natl Inst Infect Dis, Dept Virol, Tokyo 1628460, Japan

[3] Toray Industries Ltd, Pharmaceut Res Labs, Kanagawa 2488555, Japan

来源：

NATURE PROTOCOLS | 2006年 / 1卷 / 05期

基金：

日本学术振兴会;

关键词：

D O I：

10.1038/nprot.2006.395

中图分类号：

Q5 [生物化学];

学科分类号：

071010 ; 081704 ;

摘要：

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.

引用

收藏

页码：2334 / 2339

页数：6

相关论文

共 12 条

[1] Highly permissive cell lines for subgenomic and genomic hepatitis C virus RNA replication [J].

Blight, KJ ;

McKeating, JA ;

Rice, CM .

JOURNAL OF VIROLOGY, 2002, 76 (24) :13001-13014

[2] Mutations that permit efficient replication of hepatitis C virus RNA in Huh-7 cells prevent productive replication in chimpanzees [J].

Bukh, J ;

Pietschmann, T ;

Lohmann, V ;

Krieger, N ;

Faulk, K ;

Engle, RE ;

Govindarajan, S ;

Shapiro, M ;

Claire, MS ;

Bartenschlager, R .

PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 2002, 99 (22) :14416-14421

[3] Genotype 2a hepatitis C virus subgenomic replicon can replicate in HepG2 and IMY-N9 cells [J].

Date, T ;

Kato, T ;

Miyamoto, M ;

Zhao, ZJ ;

Yasui, K ;

Mizokami, M ;

Wakita, T .

JOURNAL OF BIOLOGICAL CHEMISTRY, 2004, 279 (21) :22371-22376

[4] Efficient replication of the genotype 2a hepatitis C virus subgenomic replicon [J].

Kato, T ;

Date, T ;

Miyamoto, M ;

Furusaka, A ;

Tokushige, K ;

Mizokami, M ;

Wakita, T .

GASTROENTEROLOGY, 2003, 125 (06) :1808-1817

[5] Nonhepatic cell lines HeLa and 293 support efficient replication of the hepatitis C virus genotype 2a subgenomic replicon [J].

Kato, T ;

Date, T ;

Miyamoto, M ;

Zhao, ZJ ;

Mizokami, M ;

Wakita, T .

JOURNAL OF VIROLOGY, 2005, 79 (01) :592-596

[6] Sequence analysis of hepatitis C virus isolated from a fulminant hepatitis patient [J].

Kato, T ;

Furusaka, A ;

Miyamoto, M ;

Date, T ;

Yasui, K ;

Hiramoto, J ;

Nagayama, K ;

Tanaka, T ;

Wakita, T .

JOURNAL OF MEDICAL VIROLOGY, 2001, 64 (03) :334-339

[7] Complete replication of hepatitis C virus in cell culture [J].

Lindenbach, BD ;

Evans, MJ ;

Syder, AJ ;

Wölk, B ;

Tellinghuisen, TL ;

Liu, CC ;

Maruyama, T ;

Hynes, RO ;

Burton, DR ;

McKeating, JA ;

Rice, CM .

SCIENCE, 2005, 309 (5734) :623-626

[8] Real-time detection system for quantification of hepatitis C virus genome [J].

Takeuchi, T ;

Katsume, A ;

Tanaka, T ;

Abe, A ;

Inoue, K ;

Tsukiyama-Kohara, K ;

Kawaguchi, R ;

Tanaka, S ;

Kohara, M .

GASTROENTEROLOGY, 1999, 116 (03) :636-642

[9] ELECTROPORATION IN INTRACELLULAR BUFFER INCREASES CELL-SURVIVAL [J].

VANDENHOFF, MJB ;

MOORMAN, AFM ;

LAMERS, WH .

NUCLEIC ACIDS RESEARCH, 1992, 20 (11) :2902-2902

[10] Production of infectious hepatitis C virus in tissue culture from a cloned viral genome [J].

Wakita, T ;

Pietschmann, T ;

Kato, T ;

Date, T ;

Miyamoto, M ;

Zhao, ZJ ;

Murthy, K ;

Habermann, A ;

Kräusslich, HG ;

Mizokami, M ;

Bartenschlager, R ;

Liang, TJ .

NATURE MEDICINE, 2005, 11 (07) :791-796