Self-priming of hepatitis C virus RNA

Abstract

To identify hepatitis C virus (HCV) infection, HCV genomic ribonucleic acid (RNA) can be detected using the reverse transcription-nested polymerase chain reaction (RT-nested PCR). HCV replication involves the production of a complementary, genomic-length, negative RNA strand via semiconservative RNA synthesis, utilizing negative strand specific reverse transcription and subsequent DNA synthesis. It is important to exclude the presence of self-priming in negative strand specific RT-nested PCR assay. In these experiments, HCV genomic RNA was subjected to reverse transcription without addition of a primer, and the resultant complementary deoxyribonucleic acid (cDNA) was produced. Digestion of template with RNase suggests that the template RNA is reverse transcribed with an RNA primer, not a DNA primer. Therefore, caution must be employed in interpreting studies of HCV replication using negative strand specific reverse transcription and PCR.