Abstract
The amplification mechanism of siRNAs along the transgene sequence exists in RNAi. The RNA-dependent RNA polymerase synthesizes complementary RNAs by using the transgene mRNA as a template, and the secondary siRNAs are generated from the outside of primary RNAi target. We addressed the transitive RNA silencing of a tobacco endoplasmic reticulum omega-3 fatty acid desaturase gene (NtFAD3). Four independent RNAi vectors which produced primary siRNAs against distinct regions of NtFAD3 were transiently expressed in the leaves of the NtFAD3-overexpressed transgenic plants by agroinfiltration. Irrespective of RNAi vectors used, the secondary NtFAD3 siRNAs were generated only from the 3' downstream region of the transgene but not from the 5' upstream region relative to the annealing site of primary siRNAs. These results suggest that regulatory mechanisms involved in the spreading of RNA silencing differentially act on the 5' upstream and 3' downstream regions of the target sequence, respectively.
