siPOOLs: robust reagents for gene silencing
Although we talk a lot about off-targets, one of the main advantages of siPOOLs (complex siRNA pools) compared to single siRNAs or mini-pools (Dharmacon) is that they provide near optimum silencing of target genes.
Similar seed effects in independent siRNA screens
A 2013 study on Parkin translocation used genome-wide siRNA libraries from Ambion (single Silencer Select siRNAs) and Dharmacon (pools of 4 siGENOME siRNAs).
Performing target validation well
This blogpost describes issues encountered in target validation and how to safeguard against poor reproducibility in RNAi experiments.
Low complexity pooling does not prevent siRNA off-targets
Low-complexity siRNA pooling (e.g. Dharmacon siGENOME SMARTpools) does not prevent siRNA off-targets. It may in fact exacerbate off-target effects. Only high-complexity pooling (siPOOLs) can reliably ensure on-target phenotypes.
What is the probability of an siRNA off-target phenotype?
Conventional siRNAs have a high probability of giving off-target phenotypes. siRNA off-target effects can be reduced by using more specific reagents or narrowing the assay focus (to reduce the number of relevant genes).
5 factors to consider in multi-gene targeting RNAi screens
Effective functional genomic screening depends on a variety of factors that need to be simultaneously addressed to obtain meaningful results. A recent Cell Reports paper demonstrates this by taking a holistic approach to siRNA screening with the use of multi-isoform/multi-gene targeting to address redundant paralogs and pathways in cancer cells.
Is it important to avoid microRNA binding sites during siRNA design?
To address the question of whether one should avoid microRNA binding sites during siRNA design, we examined whether removing siRNAs that share seeds with native microRNAs would reduce the dominance of seed-based off-target effects in RNAi screening.
Disrupting lncRNA function with siPOOLs (RNAi), antisense oligos and CRISPR
This blogpost covers methods used in the disruption of lncRNA function. Specifically focusing on RNA interference (with siPOOLs), antisense oligos, and CRISPR approaches. Challenges faced with these approaches are addressed.
Correcting seed-based off-target effects in RNAi screens
Unlocking RNAi screening potential by correcting seed-based off-target effects. While seed correction methods yield minor improvements, challenges persist in interpreting strongest hits. The quest for precise reagents, like high-complexity siPOOLs, remains crucial for reliable results.
Making sense of siGENOME deconvolution
Deconvolution of Dharmacon siGENOME pools shows surprising results, with low correlation to target genes and significant seed effects.