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Validation of RNAi silencing specificity using synthetic genes: salicylic acid‐binding protein 2 is required for innate immunity in plants
Author(s) -
Kumar Dhirendra,
Gustafsson Claes,
Klessig Daniel F.
Publication year - 2006
Publication title -
the plant journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.058
H-Index - 269
eISSN - 1365-313X
pISSN - 0960-7412
DOI - 10.1111/j.1365-313x.2005.02645.x
Subject(s) - rna interference , gene silencing , biology , gene , computational biology , genetics , innate immune system , microbiology and biotechnology , rna , immune system
SummaryRNA interference (RNAi) is widely used to specifically silence the expression of any gene to study its function and to identify and validate therapeutic targets. Despite the popularity of this technology, recent studies have shown that RNAi may also silence non‐targeted genes. Here we demonstrate the utility of a quick, efficient and robust approach to directly validate the specificity of RNAi as an alternative to indirect validation of RNAi through gene expression profiling. Our approach involves reversing (complementing) the RNAi‐induced phenotype by introducing a synthetic version of the target gene that is designed to escape silencing. This synthetic gene complementation approach can also be used for mutational analysis of the target gene, or to provide a functional version of a defective protein after silencing the defective gene by RNAi. Using this approach we demonstrate that the loss of systemic acquired resistance, a form of innate immunity in plants, is indeed due to the silencing of salicylic acid‐binding protein 2 rather than to off‐target effects.