Bean Pod Mottle Virus as a Powerful VIGS Vector for Functional Genomics in Pisum sativum

Pea (Pisum sativum L.) is an important agronomic crop legume worldwide. Complete sequencing of the pea genome is currently underway and will lead to the identification of many genes potentially associated with important agronomic traits. Because stable genetic transformation is laborious for pea, virus-induced gene silencing (VIGS) appears to be a powerful alternative technology for determining the function of unknown genes. In this work, we present a rapid and efficient viral inoculation method using DNA infectious plasmids of BPMV-derived VIGS vector. Six pea genotypes with important biotic and/or abiotic genes were found susceptible to BPMV carrying GFP reporter gene and showed fluorescence in shoots and roots. In a second step, we investigated 37 additional pea genotypes and found that 30 were susceptible to BPMV and only 7 were resistant. The capacity of BPMV to induce silencing of endogenes was investigated in the most susceptible genotype using two visual reporter genes: PsPDS and PsKORRIGAN1 (PsKOR1) encoding PHYTOENE DESATURASE and a 1,4-β-D-glucanase, respectively. The features of the “one-step” BPMV-derived VIGS vector include: (i) ease of rub-inoculation, without any need for biolistic or agro-inoculation procedures, (ii) simple cost-effective procedure applicable in all laboratories, (iii) non-interference of viral symptoms with silencing or over-expression phenotypes. These features make BPMV the most adapted VIGS vector in pea to make low- to high-throughput VIGS studies.

Keywords: Pisum sativum, virus-induced gene silencing, Bean pod mottle virus, PDS, KORRIGAN1