Precise genome-wide base editing by the CRISPR Nickase system in yeast.
Precise genome-wide base editing by the CRISPR Nickase system in yeast.
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DOI：
10.1038/s41598-017-02013-7
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发表时间：
2017-05-18
影响因子：
4.6
通讯作者：
Ueda M
中科院分区：
文献类型：
--
作者：
Satomura A;Nishioka R;Mori H;Sato K;Kuroda K;Ueda M
CRISPR/Cas9系统已被应用于许多真核细胞的高效基因组编辑。然而，该系统可以编辑的碱基仅限于Protspacer相邻基序(PAM)和引导RNA靶向序列中的碱基。在这项研究中，我们开发了一种全基因组碱基编辑技术，“CRISPR镍酶系统”，它利用一个单一的Cas9镍酶。该系统不受CRISPR/CAS9系统中观察到的可编辑碱基的限制，能够从划痕位置精确编辑高达53 碱基。此外，与CRISPR/CAS9系统相比，该系统没有表现出偏离目标的编辑。将CRISPR镍酶系统与酵母缺口修复克隆相结合，仅在5天内就构建了酵母突变株。CRISPR镍酶系统为酵母中快速、特定部位和精确的碱基编辑提供了一种通用和强大的技术。
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.