Development of a Full-Length Infectious Clone of Grapevine rupestris stem pitting-associated virus strain Syrah and GFP-Tagged and VIGS Vectors for Vitis vinifera

Abstract

Grapevines are an important Canadian crop, yet quality, health, and yield are threatened by viral infections and coinfections with multiple viruses complicate disease associations. Grapevine rupestris stem pitting-associated virus (GRSPaV) is a single-stranded, positive-sense RNA virus that is widely distributed worldwide and is associated with Rupestris Stem Pitting, Syrah Decline, and Grapevine Vein Necrosis. Specifically the SY strain is associated with unhealthy vines, possibly being more pathogenic. An infectious clone can help clarify disease associations and viral biology by reproducing singular infections in virus-free plants. MGT, a clone related to SY, was constructed and a GFP-tagged version produced GFP faster and to a greater degree than the asymptomatic GG strain in Nicotiana benthamiana, as determined by Western blotting and epifluorescence microscopy. A Virus-Induced Gene Silencing (VIGS) clone was also constructed for future study of grapevine genomics. These N. benthamiana experiments provide a foundation for continued work with this clone with grapevines.