RNAi off-targeting: Light at the end of the tunnel

Synthetic 19 base pair (bp) RNA duplexes (small interfering RNAs, siRNAs) can enter RISC directly and induce RNAi-mediated cleavage of targeted transcripts without additional processing. Though this ability has enormous potential, expansion of RNAi technology requires overcoming obstacles associated with siRNA functionality and specificity. Initial problems associated with inconsistent gene knockdown were found to be inherent in the sequence and thermodynamic properties of the siRNA and were addressed by development of algorithms that enabled the selection of highly functional sequences. In contrast, three separate attributes appear to contribute to overall siRNA specificity, thus making precise knockdown a surprisingly onerous problem.