Virus-Induced Gene Silencing as a Scalable Tool to Study Drought Tolerance in Plants
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Here we describe the methodology of using virus-induced gene silencing (VIGS) as a powerful and scalable tool to screen the function of genes that participate in adaptation to drought. Silencing of endogenous gene expression in Nicotiana benthamiana is achieved by systemic infection of the aerial parts of the plant with a virus engineered to contain homologous fragments of the target gene(s) of interest. Silenced plant material can be consistently produced with little optimization in less than 1 month without specialized equipment, using only simple cloning and transformation techniques. Although maximal silencing is localized to only a few leaves, when whole plants are subjected to water stress, the tissue from these silenced leaves can be characterized for physiological, biochemical, and transcriptional responses to determine the role of the candidate genes in drought tolerance.
Key wordsVirus-induced gene silencing Drought response/s Reverse genetics
We gratefully acknowledge the Swiss-South African Joint Research Program (Grant Number: S002533) for support in funding this project. In addition, we would like to thank Prof. Samuel C. Zeeman, Prof. Jens Kossmann, the Zürich-Basel Plant Science Centers’ Plant Fellows program, and the South African National Research Foundation for their continuing contributions and support.
- 22.Granier C, Aguirrezabal L, Chenu K et al (2006) PHENOPSIS, an automated platform for reproducible phenotyping of plant responses to soil water deficit in Arabidopsis thaliana permitted the identification of an accession with low sensitivity to soil water deficit. New Phytol 169:623–635CrossRefPubMedGoogle Scholar
- 26.George GM, van der Merwe MJ, Nunes-Nesi A et al (2010) Virus-induced gene silencing of plastidial soluble inorganic pyrophosphatase impairs essential leaf anabolic pathways and reduces drought stress tolerance in Nicotiana benthamiana. Plant Physiol 154:55–66CrossRefPubMedPubMedCentralGoogle Scholar
- 30.Sunkar R (2010) Plant stress tolerance: methods and protocols. Methods Mol Biol 639:386Google Scholar
- 31.Abramoff MD, Magalhaes PJ, Ram SJ (2004) Image processing with ImageJ. Biophoton Int 11:36–42Google Scholar