International Journal of Innovative Horticulture

Potato viruses including Potato virus S (PVS), Potato virus M (PVM), Potato leafroll virus (PLRV) and Potato virus X (PVX) are major viruses infecting potato and can cause serious crop losses. In this study, a multiplex reverse transcription polymerase chain reaction (RT-PCR) protocol was developed for simultaneous detection of PVS, PVM, PLRV and PVX. At least three specific primers pairs were designed for each virus within the conserved regions of coat protein gene for single and/or multiplex RT-PCR. Finally, one pair of primer for each virus was selected for multiplex RT-PCR producing distinct fragments 263, 408, 452 and 565 bp, representing PVS, PVM, PLRV and PVX, respectively. All the primers were designed based on specificity and compatibility and the amplicons were confirmed by sequencing and BLAST analysis. The sensitivity analysis of the multiplex RT-PCR was done using a 10-fold serial dilution of total RNA and protocol was sensitive enough to detect up to 10^"4 for PVM, PLRV and PVX and 10^"3 for PVS. The protocol was validated using field samples (leaves and tubers) collected from different parts of India. The simultaneous and sensitive detection of different viruses using the multiplex RT-PCR is more efficient and economical than other conventional methods for potato virus detection. The result indicates the robustness of the protocol will be useful for virus indexing of mother stocks in the seed production system and for epidemiological studies.