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siPOOL



siPOOL

Exceptional Gene Silencing Specificity For Reliable Phenotypes



For targeted gene silencing, RNA interference (RNAi) presents multiple benefits of fast results, easy setup, high efficiency and broad applicability to various cell types. Its transient effects and dose-dependent behavior closely resembles that of small molecules. Yet current RNAi reagents suffer profoundly from poor specificity and variable gene silencing efficiencies, hindering their application as a drug discovery and gene research tool.

siRNA pools (siPOOLs) are high complexity pools of 30 optimally-designed siRNAs that were demonstrated to efficiently remove off-target effects and improve reliability of results (Hannus et al., 2014). The “Pack Hunter” (pooling) approach counters off-targeting of individual siRNAs by diluting¬† their concentrations below thresholds that stimulate phenotypes. With proprietary design algorithms, siRNA sequences within siPOOLs are also optimized for maximal transcript coverage, efficient hybridization and filtered against paralogues, enabling highly efficient and specific gene silencing.

siPOOLs are suited for use against coding or long non-coding RNAs, selected isoforms or closely-related genes, and can be applied in combination to study synergistic gene interactions (Welsbie et al., 2017).

siPOOLs are ideal tools in drug discovery for target validation and target identification with custom or ready-made siPOOL libraries.




Advantages of Our Technology



01 Simple and fast to use

siPOOLs are compatible with a wide-range of transfection reagents and results are seen within days.

02 Highly specific and efficient

siPOOLs reduce off-targeting by 5-25 fold and achieve ‚Č• 70% gene knock-down at 1-3 nM in standard cell lines.

03 Consistent phenotypes

Phenotypes produced by sequence-independent siPOOLs are highly correlated as opposed to siRNAs.

04 Validation-inclusive Guarantee

siPOOL validation by rtqPCR is performed with possibility of re-design if less than 70% knock-down is seen under optimal transfection.

05 Customized design using latest annotations

Detailed design ensures optimized thermodynamics and paralog avoidance. Sequence info available.

06 PAGE-purified and non-toxic

All siPOOLs are PAGE-purified reducing risk of contaminants and side-effects.


How it works

siPOOL Technology




Individual siRNAs or low complexity siRNA pools containing 3-4 siRNAs often hit multiple off-target genes and exhibit variable target gene knock-down.

Short interfering RNA (siRNA) pools, siPOOLs are highly complex and defined pools of 30 siRNAs, each siRNA present at picomolar working concentrations. This:
  • dilutes the off-target signature of each siRNA, increasing on-target specificity.
  • ensures co-operative knock-down of the target gene, producing more robust and reliable results.

 


Optimized and Detailed siRNA Design

With defined siRNA sequences within the siPOOL, targeting can be optimized against specific transcript isoforms or closely related genes. Proprietary siRNA design algorithms select the most potent siRNAs based on thermodynamic properties that favour guide strand loading into the RNA-induced silencing complex (RISC) (refer Technote 2 for more details on siPOOL design). Using latest RefSeq annotations and genome-wide paralogue filtering, siPOOLs are designed for maximum coverage of all targeted transcripts with high specificity.



Supporting Data




Higher specificity with siPOOLs



siRNA
siRNA


Specific reagents should only affect their target.

Expression profiling by microarray in HeLa cells revealed a single siRNA can induce numerous off-target genes (red dots) while a siPOOL against the same target gene (green dot), and containing the non-specific siRNA, had greatly reduced off-target effects.



Better reproducibility and potent knock-down efficiency with siPOOLs



siRNA
siPOOL
siPOOL


siRNAs vary strongly in their knock-down efficiency.

siPOOLs against the same gene gave similar knock-down efficiencies compared to single siRNAs, indicating greater robustness and reproducibility (left and middle panel: correlation plots of real-time quantitative PCR measurements of target RNA levels).

siPOOLs also exhibit potent gene knock-down. In commonly used cell lines, gene knock-down efficiencies of 75-98% are often achieved at 1 nM concentrations for many genes (right panel).




Phenotypes you can trust with siPOOLs



siRNA
siRNA


If reliable and specific, two reagents that target the same gene should produce similar phenotypes.

We screened 36 genes using two siPOOLs per gene from our human kinase siPOOL library and three siRNAs per gene from a commercially available library and measured cell viability in A549 cells. Only siPOOLs produced consistent phenotypes.

For a complete publication on siPOOLs:

Hannus M. et. al. siPools: highly complex but accurately defined siRNA pools eliminate off-target effects. Nucleic Acids Res. (2014)



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