Separate detection of the two complementary RNA strands of hepatitis A virus.
The minus strand of hepatitis A virus can be detected specifically by reverse transcription and polymerase chain reaction amplification in infected cell culture extracts. Several controls gave evidence that the amplified fragment actually used the minus strand as initial template. Non-thermostable reverse transcriptase was not efficient for this purpose because of self-priming of the positive-stranded viral RNA during the reverse transcription step. This problem was overcome by the use of the thermostable rTth DNA polymerase that also has reverse transcriptase activity in the presence of Mn2+.
Agnès
F
F
Centre de Recherches du Service de Santé des Armées, Unité de Biologie Moléculaire, La Tronche, France.
Crance
J M
JM
Lévêque
F
F
D009189:Avian Myeloblastosis Virus / Q000201:enzymology; D001483:Base Sequence; D002460:Cell Line; D017931:DNA Primers / Q000235:genetics; D004259:DNA-Directed DNA Polymerase; D006507:Hepatovirus / Q000235:genetics* / Q000302:isolation & purification*; D006801:Humans; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods; D018075:RNA, Complementary / Q000235:genetics* / Q000302:isolation & purification*; D012367:RNA, Viral / Q000235:genetics* / Q000302:isolation & purification*; D012194:RNA-Directed DNA Polymerase; D014158:Transcription, Genetic; D014773:Virology / Q000379:methods*
D017931:DNA Primers; D018075:RNA, Complementary; D012367:RNA, Viral; C068508:Tth polymerase; D012194:RNA-Directed DNA Polymerase; D004259:DNA-Directed DNA Polymerase