Similar seed effects in independent siRNA screens

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).

The correlation between results for the same on-target gene from the two libraries was very low (R = 0.09). (Each point in the following plot is for a gene.)

% Parkin Translocation (PPT) for Ambion vs. Dharmacon siRNAs grouped by same 7mer seed

The correlation between results for the same 7mer seed were higher (0.26), providing another example of the Iron Law of RNAi Screening. (Each point in the following plot is for a 7mer seed.)

It is also worth noting that the seed-based correlation would likely have been much higher, had the Dharmacon siRNAs been screened individually (see details below).

Conclusion

The only effective way to avoid off-target effects in RNAi screening is to use high-complexity reagents like siPOOLs, which dilute away off-target effects while maintaining strong on-target silencing.

Analysis details

To calculate the Ambion by-gene value, the mean PPT value was taken for the 3 on-target siRNAs for the gene. (The Dharmacon pooled library only has 1 value per gene, so no further calculation is necessary.)

To calculate the Ambion by-seed value, the mean PPT value was taken for all siRNAs with the 7mer. For Dharmacon, the pool value was assigned to each siRNA, and then siRNAs were grouped by their 7mer seed in order to calculate the seed mean. This means that the Dharmacon siRNA seed value is actually the average from 4 different siRNAs (with different seeds). Had the Dharmacon siRNAs been screened individually, the correlation with Ambion seed results would have been higher.

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