Abstract
Several VIGS protocols have been established for high-throughput functional genomic screens as it bypasses the time-consuming and laborious process of generation of transgenic plants. The silencing efficiency in this approach is largely hindered by a technically demanding step in which the first pair of newly emerged true leaves at the 2-week-old stage are infiltrated with a needleless syringe. To further optimize VIGS efficiency and achieve rapid inoculation for a large-scale functional genomic study, here we describe a protocol of an efficient VIGS assay in Arabidopsis using Agrobacterium-mediated rubbing infection. The Agrobacterium inoculation is performed by simply rubbing the leaves with Filter Agent Celite® 545. The highly efficient and uniform silencing effect was indicated by the development of a visibly albino phenotype due to silencing of the Cloroplastos alterados 1 (CLA1) gene in the newly emerged leaves. In addition, the albino phenotype could be observed in stems and flowers, indicating its potential application for gene functional studies in the late vegetative development and flowering stages.
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Acknowledgment
We thank the support from USDA NIFA (2012-67013-19433), the Robert A. Welch Foundation (A-1795) to L. S., and the National Council for Scientific and Technological Development-Brazil (CNPq) to A. M.
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Manhães, A.M.E.d.A., de Oliveira, M.V.V., Shan, L. (2015). Establishment of an Efficient Virus-Induced Gene Silencing (VIGS) Assay in Arabidopsis by Agrobacterium-Mediated Rubbing Infection. In: Mysore, K., Senthil-Kumar, M. (eds) Plant Gene Silencing. Methods in Molecular Biology, vol 1287. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-2453-0_17
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DOI: https://doi.org/10.1007/978-1-4939-2453-0_17
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