Chu, Chia-yingRana, Tariq M.2022-08-232022-08-232008-07-292009-10-22<p>RNA. 2008 Sep;14(9):1714-9. Epub 2008 Jul 24. <a href="http://dx.doi.org/10.1261/rna.1161908">Link to article on publisher's site</a></p>1469-9001 (Electronic)10.1261/rna.116190818658119https://hdl.handle.net/20.500.14038/39105RNA interference (RNAi) is a gene-silencing mechanism by which a ribonucleoprotein complex, the RNA-induced silencing complex (RISC) and a double-stranded (ds) short-interfering RNA (siRNA), targets a complementary mRNA for site-specific cleavage and subsequent degradation. While longer dsRNA are endogenously processed into 21- to 24-nucleotide (nt) siRNAs or miRNAs to induce gene silencing, RNAi studies in human cells typically use synthetic 19- to 20-nt siRNA duplexes with 2-nt overhangs at the 3'-end of both strands. Here, we report that systematic synthesis and analysis of siRNAs with deletions at the passenger and/or guide strand revealed a short RNAi trigger, 16-nt siRNA, which induces potent RNAi in human cells. Our results indicate that the minimal requirement for dsRNA to trigger RNAi is an approximately 42 A A-form helix with approximately 1.5 helical turns. The 16-nt siRNA more effectively knocked down mRNA and protein levels than 19-nt siRNA when targeting the endogenous CDK9 gene, suggesting that 16-nt siRNA is a more potent RNAi trigger. In vitro kinetic analysis of RNA-induced silencing complex (RISC) programmed in HeLa cells indicates that 16-nt siRNA has a higher RISC-loading capacity than 19-nt siRNA. These results suggest that RISC assembly and activation during RNAi does not necessarily require a 19-nt duplex siRNA and that 16-nt duplexes can be designed as more potent triggers to induce RNAi.en-USBase SequenceHela CellsHumans*RNA InterferenceRNA, Double-StrandedRNA, Small InterferingRNA-Induced Silencing ComplexLife SciencesMedicine and Health SciencesJournal Articlehttps://escholarship.umassmed.edu/oapubs/19211042848oapubs/1921