Development of a multiplex PCR assay for simultaneous detection of monopartite and bipartite begomoviruses along with associated satellite DNAs

Begomoviral diseases are one of the major factors behind economic losses in agriculturally important crops. Begomoviruses are transmitted by whiteflies ( Bemisia tabaci ) and are often found to be associated with alphasatellites and betasatellites. Rapid, reliable and early detection methods are invaluable in developing control strategies against these viruses. A multiplex polymerase chain reaction (mPCR) assay was developed for simultaneous detection of monopartite and bipartite begomoviruses along with the associated satellites. Symptomatic leaves of different plant species were used for begomovirus and satellite detection. Primer pairs for each component were designed using reference sequences from GenBank. The primer pairs were designed in such a way that the amplicons differ at least 100bp in size. Amplicons of ~ 185bp, ~ 290bp, ~ 420bp and ~ 685bp were obtained in the mPCR that correspond to betasatellite, DNA-A, alphasatellite and DNA-B respectively. Identities of the viral and satellite components were confirmed by sequencing of the amplicons obtained in mPCR. This mPCR assay can be used for rapid and simultaneous detection of begomovirus genomic components and associated satellites.