What is it about?
RNA interference (RNAi) is a widely used technique that allows selective down regulation of genes of interest. For this purpose small interfering RNAs (siRNAs) can be transfected into cells in order to bring about sequence specific cleavage of target RNAs. Our publication underlines the necessity for carefully designed negative control siRNAs and introduces a novel class of negative control siRNA with superior properties as compared to standard controls. Our design involves randomising defined positions on complimentary siRNA strands, thereby creating large pools of randomised negative control siRNA (rnd-siRNA). The thermodynamic properties and target selectivity of rnd-siRNAs are analysed in this publication.
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Why is it important?
So far, researchers often rely on negative control siRNAs that a growing body of evidence is showing to be inadequate (e.g. scrambled controls). The major problem of those controls is caused by "miRNA-like" off-target effects (OTEs). Here we propose a novel negative control compound that has greatly reduced OTEs.
Perspectives
The use of appropriately designed controls greatly increases the quality of experiments. We think that rnd-siRNAs have a great advantage over conventional siRNA controls: they minimise sequence-specific OTEs, while maintaining some desired non-sequence specific OTEs thereby mimicking the effector siRNA. Especially large transcriptome-wide gene perturbation screens consisting of multiple siRNA treatments can profit from rnd-siRNAs, which increase the reliability of findings. Rnd-siRNAs furthermore are easily synthesized, form thermodynamically stable duplexes and consist of at least one order of magnitude larger siRNA pools than esiRNA or pooled siRNA controls.
Dr Julian A Zagalak
ETH Zürich, D-CHAB
Read the Original
This page is a summary of: Properties of short double-stranded RNAs carrying randomized base pairs: toward better controls for RNAi experiments, RNA, October 2015, Cold Spring Harbor Laboratory Press,
DOI: 10.1261/rna.053637.115.
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