Reverse transcription and subsequent DNA amplification of rubella virus RNA.
A method is described whereby rubella virus RNA was reverse transcribed and the resulting cDNA enzymatically amplified using Taq polymerase. The reactions were carried out in a single reaction vessel, with only minor modifications to the buffer conditions between the reverse transcription and the subsequent amplification step. Using an oligonucleotide probe to the E1 glycoprotein region and limited restriction endonuclease mapping, the resulting amplified products were shown to be specific for rubella virus. This method was also successfully applied to crude cell lysates, without the need for RNA purification. The possible applications of the polymerase chain reaction as applied to RNA sequences are discussed.
Carman
W F
WF
National Institute for Virology, University of the Witwatersrand, Johannesburg, South Africa.
Williamson
C
C
Cunliffe
B A
BA
Kidd
A H
AH
D000818:Animals; D002522:Chlorocebus aethiops; D004279:DNA, Viral / Q000302:isolation & purification; D004259:DNA-Directed DNA Polymerase; D005784:Gene Amplification; D006678:HIV / Q000235:genetics; D006801:Humans; D015345:Oligonucleotide Probes; D012367:RNA, Viral / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism*; D012194:RNA-Directed DNA Polymerase; D012412:Rubella virus / Q000235:genetics*; D019914:Taq Polymerase; D014158:Transcription, Genetic; D014709:Vero Cells
D004279:DNA, Viral; D015345:Oligonucleotide Probes; D012367:RNA, Viral; D019914:Taq Polymerase; D012194:RNA-Directed DNA Polymerase; D004259:DNA-Directed DNA Polymerase