Development of a Reverse Genetic System for Studying Rose Rosette Virus in Whole Plants. Academic Article uri icon

abstract

  • Rose rosette virus (RRV) is a negative-sense RNA virus with a seven-segmented genome that is enclosed by a double membrane. We constructed an unconventional minireplicon system encoding the antigenomic (ag)RNA1 (encoding the viral RNA-dependent RNA polymerase [RdRp]), agRNA3 (encoding the nucleocapsid protein [N]), and a modified agRNA5 containing the coding sequence for the iLOV protein in place of the P5 open reading frame (R5-iLOV). iLOV expression from the R5-iLOV template was amplified by activities of the RdRp and N proteins in Nicotiana benthamiana leaves. A mutation was introduced into the RdRp catalytic domain and iLOV expression was eliminated, indicating RNA1-encoded polymerase activity drives iLOV expression from the R5-iLOV template. Fluorescence from the replicon was highest at 3 days postinoculation (dpi) and declined at 7 and 13 dpi. Addition of the tomato bushy stunt virus (TBSV) P19 silencing-suppressor protein prolonged expression until 7 dpi. A full-length infectious clone system was constructed of seven binary plasmids encoding each of the seven genome segments. Agro-delivery of constructs encoding RRV RNAs 1 through 4 or RNAs 1 through 7 to N. benthamiana plants produced systemic infection. Finally, agro-delivery of the full-length RRV infectious clone including all segments produced systemic infection within 60 dpi. This advance opens new opportunities for studying RRV infection biology.

published proceedings

  • Mol Plant Microbe Interact

altmetric score

  • 1.5

author list (cited authors)

  • Verchot, J., Herath, V., Urrutia, C. D., Gayral, M., Lyle, K., Shires, M. K., Ong, K., & Byrne, D.

citation count

  • 13

complete list of authors

  • Verchot, Jeanmarie||Herath, Venura||Urrutia, Cesar D||Gayral, Mathieu||Lyle, Kelsey||Shires, Madalyn K||Ong, Kevin||Byrne, David

publication date

  • August 2020