Detection of multiple potato viruses using an oligo(dT) as a common cDNA primer in multiplex RT-PCR.
A novel usage of multiplex reverse transcription polymerase chain reaction (m-RT-PCR) for simultaneous detection of multiple viruses is reported. By use of an oligo(dT), as a common primer, nearly full-length cDNAs can be synthesized. Furthermore, combining an oligo(dT) primer with a specific antisense primer can be used to simultaneously prime reverse transcription of both polyadenylated and non-polyadenylated RNAs. Four viral genera including five potato viruses [(carlavirus (PVS), polerovirus (PLRV), potexvirus (PVX), potyvirus (PVA and PVY))] and a viroid genus including a viroid genome (pospiviroid (PSTVd)) were used to develop various formats of m-RT-PCR. In artificially created viral RNA mixtures, all six RNA pathogens were detected successfully by uniplex- and m-RT-PCR. In naturally infected field grown tubers, m-RT-PCR detected infection of two to three viruses, which were present in the tubers.
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Agriculture and Agri-Food Canada, Potato Research Centre, PO Box 20280, Fredericton, New Brunswick, Canada.
Singh
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D018076:DNA, Complementary; D009838:Oligodeoxyribonucleotides; D010935:Plant Diseases / Q000821:virology; D010942:Plant Viruses / Q000235:genetics / Q000302:isolation & purification*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D011198:Solanum tuberosum / Q000821:virology*
D018076:DNA, Complementary; D009838:Oligodeoxyribonucleotides; C027903:oligo (dT)