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タイトル: Site-specific randomization of the endogenous genome by a regulatable CRISPR-Cas9 piggyBac system in human cells
著者: Ishida, Kentaro
Xu, Huaigeng
Sasakawa, Noriko
Lung, Yu, Siu, Mandy
Kudryashev, Alexandra, Julia
Gee, Peter
Hotta, Akitsu  kyouindb  KAKEN_id
著者名の別形: 石田, 健太郎
徐, 淮耕
笹川, 典子
堀田, 秋津
発行日: 10-Jan-2018
出版者: Springer Nature
誌名: Scientific Reports
巻: 8
論文番号: 310
抄録: Randomized mutagenesis at an endogenous chromosomal locus is a promising approach for protein engineering, functional assessment of regulatory elements, and modeling genetic variations. In mammalian cells, however, it is challenging to perform site-specific single-nucleotide substitution with single-stranded oligodeoxynucleotide (ssODN) donor templates due to insufficient homologous recombination and the infeasibility of positive selection. Here, we developed a DNA transposon based CRISPR-Cas9 regulated transcription and nuclear shuttling (CRONUS) system that enables the stable transduction of CRISPR-Cas9/sgRNA in broad cell types, but avoids undesired genome cleavage in the absence two chemical inducing molecules. Highly efficient single nucleotide alterations induced randomization of desired codons (up to 4 codons) at a defined genomic locus in various human cell lines, including human iPS cells. Thus, CRONUS provides a novel platform for modeling diseases and genetic variations.
著作権等: © The Author(s) 2017. This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.
URI: http://hdl.handle.net/2433/234561
DOI(出版社版): 10.1038/s41598-017-18568-4
PubMed ID: 29321585
出現コレクション:学術雑誌掲載論文等

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