Construction of mismatched inverted repeat (IR) silencing vectors for maximizing IR stability and effective gene silencing in plants

Inverted repeat (IR) RNA silencing vectors containing homologous fragments of target endogenous plant genes, or pathogen genes, are the most widely used vectors to either study the function of genes involved in biotic stress or silence pathogens to induce plant resistance, respectively. RNA silencing has been exploited to produce transgenic plants with resistance to viral pathogens via posttranscriptional gene silencing (PTGS). In some cases, this technology is difficult to apply due to the instability of IR constructs during cloning and plant transformation. We have therefore developed a robust method for the production of long IR vector constructs by introducing base pair mismatches in the form of cytosine to thymine mutations on the sense arm by exposure to sodium bisulfite prior to assembly of the IR.