CONSISTENT GENE SILENCING IN TRANSGENIC PLANTS EXPRESSING A REPLICATING POTATO-VIRUS-X RNA

Tobacco plants were transformed with constructs in which the transgene was a cDNA of replicating potato virus X (PVX) RNA. The constructs, r eferred to here as amplicons, were the intact genome of PVX and PVX co nstructs modified to carry the beta-glucuronidiase (GUS) reporter gene either as an additional gene or as a replacement for the coat protein gene (PVX/GUS/CP and PVX/GUS respectively). Transformed plants carryi ng these constructs displayed several phenotypes that we attribute to post-transcriptional gene silencing. These phenotypes include the abse nce of viral symptoms, low accumulation of transgene-derived RNA, extr eme strain-specific resistance against PVX, low and non-uniform GUS ex pression (in the PVX/GUS and PVX/GUS/CP plants) and suppression of tra nsiently expressed RNA sharing homology with the transgene. Importantl y, the amplicon-mediated gene silencing was exhibited in all lines tes ted. There was no evidence of gene silencing in seven lines expressing a PVX RNA that was unable to replicate. From these data we conclude t hat the replicating viral RNA is a potent trigger of gene silencing. M oreover, amplicon-mediated gene silencing provides an important new st rategy for the consistent activation of gene silencing in transgenic p lants.