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We have developed a reliable internally controlled RT-nested PCR method for the detection of hepatitis C virus (HCV) RNA using in vitro synthesized Renilla luciferase (Rluc) RNA as an internal control. Using this method, the 5'-noncoding region of HCV RNA (144 nucleotides) and Rluc RNA (276 nucleotides) were efficiently amplified in a single tube, and the sensitivity and specificity of this method were comparable to standard RT-nested PCR. This method was successfully performed on RNA specimens obtained from in vitro HCV-infected human hepatocyte PH5CH8 cells, which support HCV replication. In addition, we demonstrated that this method was useful for the evaluation of antiviral reagents by confirming the anti-HCV activity of bovine lactoferrin, which we previously found to be a new inhibitor of HCV infection. Therefore, this method may be useful for the studies of not only HCV but also of other viruses.
Hepatitis C virus
Reverse transcriptionnested PCR (RT-nested PCR)
Acta Medica Okayama
Okayama University Medical School
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