@article{057f2cfb423f44e9ac004799b233e6a3,
title = "Cloning-free CRISPR/Cas system facilitates functional cassette knock-in in mice",
abstract = "Although the CRISPR/Cas system has enabled one-step generation of knockout mice, low success rates of cassette knock-in limit its application range. Here we show that cloning-free, direct nuclear delivery of Cas9 protein complex with chemically synthesized dual RNAs enables highly efficient target digestion, leading to generation of knock-in mice carrying a functional cassette with up to 50% efficiency, compared with just 10% by a commonly used method consisting of Cas9 mRNA and single guide RNA. Our cloning-free CRISPR/Cas system facilitates rapid one-step generation of cassette knock-in mice, accelerating functional genomic research by providing various in vivo genetic tools.",
author = "Tomomi Aida and Keiho Chiyo and Takako Usami and Harumi Ishikubo and Risa Imahashi and Yusaku Wada and Tanaka, {Kenji F.} and Tetsushi Sakuma and Takashi Yamamoto and Kohichi Tanaka",
note = "Funding Information: This work was supported in part by the Strategic Research Program for Brain Sciences to KT and Grant-in-Aid for Science Research to TA from Ministry of Education, Culture, Sports, Science and Technology of Japan, and grants to TA from MRI of TMDU, and from the president of TMDU, and the joint Usage/Research Program to KFT from MRI of TMDU. We thank A Yamanaka (Nagoya University), F Zhang (MIT), H Aizawa, K Sugiyama, T Adachi, T Nakamura, and T Hayata (MRI, TMDU), and K Watase (CBIR, TMDU) for providing plasmids and technical support. We also thank O Nureki (University of Tokyo) and JA Doudna (UC Berkeley) for their useful discussions. Publisher Copyright: {\textcopyright} 2015 Aida et al.; licensee BioMed Central.",
year = "2015",
month = apr,
day = "29",
doi = "10.1186/s13059-015-0653-x",
language = "English",
volume = "16",
journal = "Genome Biology",
issn = "1474-7596",
publisher = "BioMed Central",
number = "1",
}