Book contents
- Frontmatter
- Contents
- Preface
- Foreword
- List of abbreviations
- Part I Basic principles
- Part II Recent advances
- Part III Experimental approaches
- Part IV Protocols and techniques
- 18 Sample collection, preparation, and storage
- 19 Hepatitis C RNA in blood and tissue samples
- 20 Hepatitis C genotypes and quasispecies
- 21 Replication of hepatitis C in tissue culture
- 22 Enzyme activities of nonstructural protein 3
- 23 Characterization of nonstructural protein 5B
- Part V Some outstanding questions and emerging areas for investigation
- References
- Index
21 - Replication of hepatitis C in tissue culture
Published online by Cambridge University Press: 27 August 2009
- Frontmatter
- Contents
- Preface
- Foreword
- List of abbreviations
- Part I Basic principles
- Part II Recent advances
- Part III Experimental approaches
- Part IV Protocols and techniques
- 18 Sample collection, preparation, and storage
- 19 Hepatitis C RNA in blood and tissue samples
- 20 Hepatitis C genotypes and quasispecies
- 21 Replication of hepatitis C in tissue culture
- 22 Enzyme activities of nonstructural protein 3
- 23 Characterization of nonstructural protein 5B
- Part V Some outstanding questions and emerging areas for investigation
- References
- Index
Summary
The establishment of tissue culture systems that consistently support high levels of HCV replication is a priority not only for elucidating the replication cycle of the virus but also for screening of putative antiviral compounds. Although many groups have reported virus production in various tissue culture systems (Table 11.1), the low levels of virus produced over time has necessitated using RT/PCR for detection (Ch. 12). Given that HCV is closely related to flaviviruses (Miller & Purcell, 1990) (Table 2.1), it has been assumed that the replication cycle of HCV, like that of flaviviruses, would contain a minus strand intermediate (Chambers et al., 1990; Monath, 1990). The finding of HCV minus strand RNA in tissue culture cells (Table 11.1) and in liver samples (Fong et al., 1991b; Gerber et al., 1992; Negro et al., 1998) from infected patients is consistent with a role for minus strand in the replication cycle, although this has yet to be formally proven. From a technical perspective, detection of minus strand RNA is problematic at best, since it involves using strand-specific primers, which may also randomly prime on the viral plus strand (or on cellular mRNAs), resulting in false-positive results. In tissue culture systems that depend upon infection of susceptible cells with virus from an infectious serum, it is not always clear whether the RT/PCR results reflect de novo production of RNA and/or detection of RNA extracted from virus particles that simply adhered to the tissue culture cells.
- Type
- Chapter
- Information
- Hepatitis C VirusFrom Laboratory to Clinic, pp. 151 - 152Publisher: Cambridge University PressPrint publication year: 2002