Hwang, Woong Y.Fu, YanfangReyon, DeepakMaeder, Morgan L.Kaini, PrakritiSander, Jeffry D.Joung, J. KeithPeterson, RandallYeh, Jing-Ruey2014-02-182013Hwang, Woong Y., Yanfang Fu, Deepak Reyon, Morgan L. Maeder, Prakriti Kaini, Jeffry D. Sander, J. Keith Joung, Randall T. Peterson, and Jing-Ruey Joanna Yeh. 2013. “Heritable and Precise Zebrafish Genome Editing Using a CRISPR-Cas System.” PLoS ONE 8 (7): e68708. doi:10.1371/journal.pone.0068708. http://dx.doi.org/10.1371/journal.pone.0068708.1932-6203http://nrs.harvard.edu/urn-3:HUL.InstRepos:11717568We have previously reported a simple and customizable CRISPR (clustered regularly interspaced short palindromic repeats) RNA-guided Cas9 nuclease (RGN) system that can be used to efficiently and robustly introduce somatic indel mutations in endogenous zebrafish genes. Here we demonstrate that RGN-induced mutations are heritable, with efficiencies of germline transmission reaching as high as 100%. In addition, we extend the power of the RGN system by showing that these nucleases can be used with single-stranded oligodeoxynucleotides (ssODNs) to create precise intended sequence modifications, including single nucleotide substitutions. Finally, we describe and validate simple strategies that improve the targeting range of RGNs from 1 in every 128 basepairs (bps) of random DNA sequence to 1 in every 8 bps. Together, these advances expand the utility of the CRISPR-Cas system in the zebrafish beyond somatic indel formation to heritable and precise genome modifications.en-USBiologyBiotechnologyBioengineeringBiological Systems EngineeringGenetic EngineeringTransgenicsGeneticsGenetic MutationHeredityModel OrganismsAnimal ModelsZebrafishEngineeringJournal Article2014-02-1810.1371/journal.pone.0068708