SELF-PRIMING OF HEPATITIS-C VIRUS-RNA

To identify hepatitis C virus (HCV) infection, HCV genomic ribonucleic acid (RNA) can be detected using the reverse transcription-nested pol ymerase chain reaction (RT-nested PCR). HCV replication involves the p roduction of a complementary, genomic-length, negative RNA strand via semiconservative RNA synthesis, utilizing negative strand specific rev erse transcription and subsequent DNA synthesis. It is important to ex clude the presence of self-priming in negative strand specific RT-nest ed PCR assay. In these experiments, HCV genomic RNA was subjected to r everse transcription without addition of a primer, and the resultant c omplementary deoxyribonucleic acid (cDNA) was produced. Digestion of t emplate with RNase suggests that the template RNA is reverse transcrib ed with an RNA primer, not a DNA primer. Therefore, caution must be em ployed in interpreting studies of HCV replication using negative stran d specific reverse transcription and PCR.