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Title: Agrobacterium-mediated infection of whole plants by yellow dwarf viruses

item YOON, JU-YEON - Seoul Women'S University
item CHOI, SEUNG-KOOK - National Horticultural Research Institute - Korea
item PALUKAITIS, PETER - Seoul Women'S University
item Gray, Stewart

Submitted to: Virus Research
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 8/15/2011
Publication Date: 9/15/2011
Citation: Yoon, J., Choi, S., Palukaitis, P., Gray, S.M. 2011. Agrobacterium-mediated infection of whole plants by yellow dwarf viruses. Virus Research. 160:428-434.

Interpretive Summary: The yellow dwarf viruses affect all major cereal crops and are responsible for billions of dollars in yield reduction annually. Despite intensive effort, effective and grower accepted resistance has not been identified or developed in most cereal crops. These viruses have remained recalcitrant to most modern biological techniques for manipulating the virus genome to better understand the molecular mechanisms controlling disease development. Our work has developed the first methods to introduce infectious cloned virus molecules into whole plants. This allows scientists to make genetic modifications to the virus and introduce those modified viruses directly into plants to study how the genetic changes affect various aspects of the virus life cycle, disease development and transmission of the virus between plant hosts. The new technologies reported in this manuscript will facilitate the development of new strategies to control these viruses and reduce disease impacts on cereal crops.

Technical Abstract: Barley yellow dwarf virus-PAV (BYDV-PAV) and cereal yellow dwarf virus-RPV (CYDV-RPV) are only transmitted between host plants by aphid vectors and not by mechanical transmission. This presents a severe limitation for the use of a reverse genetics approach to analyze the effects of mutations in these viruses on plant infection and aphid transmission. Here we describe the use of agroinfection to infect plants with BYDV-PAV and CYDV-RPV. The cDNAs corresponding to the complete RNA genomes of BYDV-PAV and CYDV-RPV were cloned into a binary vector under the control of the cauliflower mosaic virus 35S promoter and the nopaline synthase transcription termination signal. The self-cleaving ribozyme from hepatitis virus D was included to produce a transcript in planta with a 3' terminus almost identical to the natural viral RNA. ELISA and RT-PCR analysis showed that the replicons of BYDV-PAV and CYDV-RPV introduced by Agrobacterium into Nicotiana benthamiana and N. clevelandii gave rise to a local infection in the infiltrated mesophyll cells. After several weeks systemic infection of phloem tissue was detected, although no systemic symptoms were observed. Three heterologous virus silencing suppressors increased the efficiency of agroinfection and accumulation of BYDV-PAV and CYDV-RPV in the two Nicotiana species. The progeny viruses purified from infiltrated tissues were successfully transmitted to oat plants by aphids, and typical yellow dwarf symptoms were observed. This study reports the first agroinfection of eudicot plants using BYDV-PAV and CYDV-RPV.