Improved PCR Detection of Blackcurrant reversion virus in Ribes and Further Evidence that It Is the Causal Agent of Reversion Disease

Abstract

Within 5 years of mechanically inoculating blackcurrant cultivars with partially purified preparations of particles of Blackcurrant reversion virus (BRV), infected plants developed leaf and flower bud symptoms typical of reversion disease, demonstrating that BRV is the causal agent of this disease. To improve the erratic immunocapture reverse transcriptase-polymerase chain reaction (RT-PCR) detection of BRV in Ribes plants, various stepwise changes were made to the original protocol. Significant improvement in the reliability and sensitivity of BRV detection was made by extracting RNA from trapped BRV particles using Triton-X 100, the design of new primers with higher annealing temperatures, and the use of 'Ready-to-go' RT-PCR beads. These features, combined with other minor changes to the protocol, improved BRV detection in reverted blackcurrant plants from <50% to >90% but the reliability of BRV detection in red currant was always very much less and was possible only using nested PCR that was developed for this purpose.