DETECTION OF 5 SEED-BORNE LEGUME VIRUSES IN ONE SENSITIVE MULTIPLEX POLYMERASE CHAIN-REACTION TEST

developed that can simultaneously test a sample, in one tube, for the presence of five seedborne legume viruses that are of great concern to legume germ plasm banks: alfalfa mosaic alfamovirus, bean yellow mosa ic potyvirus, clover yellow vein potyvirus, cucumber mosaic cucumoviru s, and subterranean clover mottle sobemovirus. RT-PCR assays were also developed for the detection of subterranean clover redleaf luteovirus and subterranean clover stunt virus, thus providing a detection syste m for all well-characterized viruses known to infect subterranean clov er. Primers were designed so that the size of the RT-PCR product was i ndicative of the virus amplified and, where sequences of more than one strain of virus were available, conserved regions were given preferen ce as primer targets. The RT-PCR assay detected all isolates tested fo r each virus and was up to five orders of magnitude more sensitive tha n ELISA.