PT - JOURNAL ARTICLE AU - Namita Bisaria AU - Inga Jarmoskaite AU - Daniel Herschlag TI - Specificity Principles in RNA-Guided Targeting AID - 10.1101/055954 DP - 2016 Jan 01 TA - bioRxiv PG - 055954 4099 - http://biorxiv.org/content/early/2016/05/28/055954.short 4100 - http://biorxiv.org/content/early/2016/05/28/055954.full AB - RNA-guided nucleases (RGNs) provide sequence-specific gene regulation through base-pairing interactions between a small RNA guide and target RNA or DNA. RGN systems, which include CRISPR-Cas9 and RNA interference (RNAi), hold tremendous promise as programmable tools for engineering and therapeutic purposes. However, pervasive targeting of sequences that closely resemble the intended target has remained a major challenge, limiting the reliability and interpretation of RGN activity and the range of possible applications. Efforts to reduce off-target activity and enhance RGN specificity have led to a collection of empirically derived rules, which often paradoxically include decreased binding affinity of the RNA-guided nuclease to its target. Here we demonstrate that simple kinetic considerations of the targeting reaction can explain these and other literature observations. The kinetic models described provide a foundation for understanding RGN systems and a necessary physical and functional framework for their rational engineering.