Precise genome-wide base editing by the CRISPR Nickase system in yeast

Scientific Reports
Atsushi SatomuraMitsuyoshi Ueda

Abstract

The CRISPR/Cas9 system has been applied to efficient genome editing in many eukaryotic cells. However, the bases that can be edited by this system have been limited to those within the protospacer adjacent motif (PAM) and guide RNA-targeting sequences. In this study, we developed a genome-wide base editing technology, "CRISPR Nickase system" that utilizes a single Cas9 nickase. This system was free from the limitation of editable bases that was observed in the CRISPR/Cas9 system, and was able to precisely edit bases up to 53 bp from the nicking site. In addition, this system showed no off-target editing, in contrast to the CRISPR/Cas9 system. Coupling the CRISPR Nickase system with yeast gap repair cloning enabled the construction of yeast mutants within only five days. The CRISPR Nickase system provides a versatile and powerful technology for rapid, site-specific, and precise base editing in yeast.

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Citations

Apr 8, 2018·Current Genetics·Inga SoreanuAmir Aharoni
Jan 14, 2018·Applied and Environmental Microbiology·Kaifeng LiShouwen Chen
Sep 2, 2018·Medicinal Research Reviews·Wenyi WuHetian Lei
Oct 14, 2017·RNA Biology·Jie YangZhaohui Gong
Mar 6, 2019·Nature Microbiology·Becky Xu Hua FuAndrew Z Fire
Nov 7, 2020·Development, Growth & Differentiation·Yu MurakamiMasato Kinoshita
Dec 1, 2020·Frontiers in Cellular and Infection Microbiology·Prashant KumarKuldeep Dhama
Mar 29, 2021·Methods : a Companion to Methods in Enzymology·Jiajie KuangJun Zhao
Feb 14, 2019·G3 : Genes - Genomes - Genetics·Aki Hayashi, Katsunori Tanaka
Jan 9, 2019·Biochemistry·Zihe LiuJens Nielsen
Oct 2, 2021·Scientific Reports·Mollie S SchubertGarrett R Rettig
Sep 13, 2020·Journal of Microbiological Methods·Dong Xuan NguyenYoichi Honda

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Methods Mentioned

BETA
gene knockout
PCR
nucleotide exchange
flow cytometry

Software Mentioned

TALEN
Genome Analysis Toolkit
AppSan

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