Analysis of transitive RNA silencing after grafting in transgenic plants with the coat protein gene of Sweet potato feathery mottle virus.

We have previously reported the graft transmission of target specificity for RNA silencing using transgenic Nicotiana benthamiana plants expressing the coat protein gene (CP, including the 3' non-translated region) of Sweet potato feathery mottle virus. Transgenic plants carrying the 5' 200 and 400 bp regions of CP were newly produced. From these plants, two silenced and two non-silenced lines were selected to investigate the manifestation of transitive RNA silencing by graft experiments. Non-silenced scions carrying the entire transgene were grafted onto either 5' or 3' silencing inducer rootstocks. When non-silenced scions were grafted onto 5' silencing inducer rootstocks, RNA silencing was induced in the non-silenced scions and spread toward the 3' region of the transgene mRNA. Similarly, when non-silenced scions were grafted onto 3' silencing inducer rootstocks, RNA silencing was induced in the non-silenced scions, but was restricted to the 3' region of the transgene and did not spread to the 5' region. In addition, results from crossing experiments, involving non-silenced and 3' silencing inducer plants, confirmed the above finding. This indicates that RNA silencing spreads in the 5'-3' direction, not in the 3'-5' direction, along the transgene mRNA.