RT Journal Article
SR Electronic
T1 Precise Cas9 targeting enables genomic mutation prevention
JF bioRxiv
FD Cold Spring Harbor Laboratory
SP 058974
DO 10.1101/058974
A1 Chavez, Alejandro
A1 Pruitt, Benjamin W
A1 Tuttle, Marcelle
A1 Shapiro, Rebecca S
A1 Cecchi, Ryan J
A1 Winston, Jordan
A1 Turczyk, Brian M
A1 Tung, Michael
A1 Collins, James J
A1 Church, George M
YR 2016
UL http://biorxiv.org/content/early/2016/06/14/058974.abstract
AB Here we present a generalized method of guide RNA “tuning” that enables Cas9 to discriminate between two target sites that differ by a single nucleotide polymorphism. We employ our methodology to generate a novel in vivo mutation prevention system in which Cas9 actively restricts the occurrence of undesired gain-of-function mutations within a population of engineered organisms. We further demonstrate that the system is scalable to a multitude of targets and that the general tuning and prevention concepts are portable across engineered Cas9 variants and Cas9 orthologs. Finally, we show that the designed mutation prevention system maintains robust activity even when placed within the complex environment of the mouse gastrointestinal tract.