Development of a VIGS vector based on the β-satellite DNA associated with bhendi yellow vein mosaic virus

Abstract

Bhendi yellow vein mosaic virus (BYMV) is a monopartite begomovirus with an associated β-satellite. βC1 ORF encoded by the β-satellite is the symptom determinant and a strong suppressor of post transcriptional gene silencing. To create a virus induced gene silencing vector based upon the β-satellite associated with BYVMV the βC1 ORF was replaced with multiple cloning sites. GFP transgene and plant endogenous genes Su, PDS, PCNA and AGO1 were cloned into β-satellite based VIGS vector. GFP expression was silenced in the GFP expressing transgenic 16c Nicotiana benthamiana plants infiltrated with VIGS vector carrying GFP gene inside. N. benthamiana plants infiltrated with the VIGS vector harboring the endogenous genes Su, PDS, PCNA and AGO1 produced the phenotypic symptoms yellowing of the veins, photobleaching of the veins, stunting of the plant and upward leaf curling, respectively. Real time PCR analyses revealed a reduction in the levels of the corresponding transgene or endogenous target mRNA. The β-satellite based VIGS vector was able to silence the target genes effectively. Hence, BYVMV β-satellite based VIGS vector can be used in functional genomics studies.