Screening Target Specificity of siRNAs by Rapid Amplification of cDNA Ends (RACE) for Non-Sequenced Species
- 143 Downloads
RNA interference (RNAi) is the process of sequence-specific posttranslational gene silencing triggered by double-stranded RNAs (dsRNAs). RNAi is a widely used approach for studying gene function. However, studies have shown that using siRNA can lead to off-target effects when the siRNA contains sufficient sequence identity to non-target mRNA sequences. One of the important steps in designing dsRNA is verification that it has sequence identity to only the target mRNA. In this report, we propose an approach for primary screening dsRNAs for potential off-target effects by using rapid amplification of cDNA ends. This method can be especially useful for model systems using species that have limited availability of sequence data.
KeywordssiRNA dsRNA Rapid Amplification of cDNA Ends (RACE)
We thank Drs. Zhaoqing Zheng for contributing the specimens of turtle brain and Pat Ronan for helpful comments on the manuscript. This study is supported by NIH grants NS051187 and P20 RR015567 which is designated as a Center of Biomedical Research Excellence (COBRE) to J.K.
- Soutschek J, Akinc A, Bramlage B, Charisse K, Constien R, Donoghue M, Elbashir S, Geick A, Hadwiger P, Harborth J, John M, Kesavan V, Lavine G, Pandey RK, Racie T, Rajeev KG, Röhl I, Toudjarska I, Wang G, Wuschko S, Bumcrot D, Koteliansky V, Limmer S, Manoharan M, Vornlocher HP (2004) Therapeutic silencing of an endogenous gene by systemic administration of modified siRNAs. Nature 432:173–178PubMedCrossRefGoogle Scholar