Comparison of genus-specific primers in RT-PCR for the broad-spectrum detection of viruses in the genus Potyvirus by plant diagnostic laboratories

• Published in: Journal of virological methods 2018-08, Vol.258, p.29-34
• Main Authors: Arous, S., Harmon, C.L., Capobianco, H.M., Polston, J.E.
• Format: Article
• Language: English
• Subjects: Degenerate primers; Diagnostics; gels; plant viruses; Potyvirus; reverse transcriptase polymerase chain reaction; RT-PCR; taxonomy; tissues; viruses
• ISSN: 0166-0934; 1879-0984
• DOI: 10.1016/j.jviromet.2018.05.003

•Twelve primer pairs were evaluated for RT-PCR detection of nine potyviruses.•Primer pair CIFor/CIRev performed the best under the assay conditions.•CIFor/CIRev is recommended for use by plant diagnostic laboratories. The Potyvirus genus is one of the largest genera of plant viruses and encompasses many economically important pathogens. While a number of degenerate primers for use in broad spectrum RT-PCR assays have been published, it is not clear which of these primers would be the most useful for use by plant diagnostic laboratories. Twelve sets of primers were tested for their ability to detect nine potyviruses in a two-step RT-PCR. Viruses were extracted from different host backgrounds and were selected to represent eight clades plus one species between clades (sensu Gibbs and Ohshima, 2010). Results of this study indicated that the primers CIFor/CIRev produced easily detectable amplicons from all nine potyviruses without non-specific amplification, false positives, or false negatives. CIFor/CIRev produced single amplicons from potyvirus-infected tissues which could be sequenced directly without gel purification to identify the virus to species.