Rapid detection of GB virus C RNA by reverse transcription-polymerase chain reaction (RT-PCR) using primers derived from the 5'nontranslated region.
A simple reverse transcription-polymerase chain reaction (RT-PCR) procedure for the detection of GB virus C (GBV-C) RNA in serum or plasma is described. In this method, total nucleic acid, extracted from a small volume of human plasma, is reverse transcribed using random hexamers. An aliquot of cDNA is then utilized in PCR employing GBV-C specific primers designed to highly conserved regions of the 5'nontranslated region (NTR). For additional sensitivity, a second round of nested amplification is performed. Reactions are analyzed on an agarose gel and samples showing an ethidium bromide stained band of the appropriate size in the first and second amplification, or in the second amplification only, are designated to be positive. This protocol allows for the rapid and sensitive detection of GBV-C infection in human plasma or serum.
Erker
J C
JC
Virus Discovery Group, Experimental Biology Research, Abbott Laboratories, North Chicago, IL 60064, USA. james.erker@add.ssw.abbott.com
Desai
S M
SM
Mushahwar
I K
IK
D017931:DNA Primers; D004587:Electrophoresis, Agar Gel; D004996:Ethidium; D018067:Flaviviridae / Q000235:genetics / Q000302:isolation & purification*; D006525:Hepatitis, Viral, Human / Q000175:diagnosis* / Q000821:virology; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods*; D012367:RNA, Viral / Q000097:blood*; D012680:Sensitivity and Specificity; D014158:Transcription, Genetic; D014766:Viremia / Q000175:diagnosis
D017931:DNA Primers; D012367:RNA, Viral; D004996:Ethidium