Emergence of mutation clusters in the HCV genome during sequential viral passages in Sip-L expressing cells

Der Ren Hwang, Hsin Yu Lai, Ming Ling Chang, John T.A. Hsu, Chau Ting Yeh*

*Corresponding author for this work

Research output: Contribution to journalJournal Article peer-review

6 Scopus citations

Abstract

Sip-L, a member of the Cupin superfamily, is a hepatic factor capable of supporting hepatitis C virus (HCV) replication in an otherwise non-permissive cell line. HCV-positive serum was used to infect Huh-7 and 293 cells stably expressing Sip-L. Using the culture medium of the infected cells as an infection source, sequential viral passages were carried out in both cell lines. Efficient viral passage was observed in 293-Sip-L cells but not in Huh-7-Sip-L cells. The viral concentrations in the culture medium increased gradually from less than 102 copies/mL to 5.3 × 104 copies/mL after 25 sequential passages in 293-Sip-L cells. Sequence analysis of the viral genomes obtained from both the initial and final inocula revealed emergence of mutation clusters in NS2, NS3, and NS5A coding regions. Immunofluorescence study revealed that only a small percentage of infected cells expressed a detectable level of viral protein. Caspase 3 activities in the infected cells increased progressively during the viral passages. In conclusion, perpetual propagation of HCV was achieved using Sip-L expressing cells, allowing for the development of mutation clusters in the genome. The mutant HCV can be used as an infection source to study the molecular mechanism of HCV replication.

Original languageEnglish
Pages (from-to)170-177
Number of pages8
JournalJournal of Virological Methods
Volume129
Issue number2
DOIs
StatePublished - 11 2005
Externally publishedYes

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