Engineered Staphylococcus auricularis Cas9 with high-fidelity

文献类型： 外文期刊

第一作者： Wei, Nan

作者： Wei, Nan;Shang, Lu;Liu, Jing;Wang, Mi;Liu, Yingchun;Zhu, Chuangang;Fei, Chenzhong;Zhang, Lifang;Yang, Fayu;Gu, Feng;Wei, Nan;Shang, Lu;Liu, Jing;Wang, Mi;Liu, Yingchun;Fei, Chenzhong;Zhang, Lifang;Yang, Fayu;Gu, Feng;Yang, Fayu;Gu, Feng

作者机构：

关键词： CRISPR-Cas9; high-fidelity; off-target effects; Staphylococcus auricularis Cas9 (SauriCas9)

期刊名称：FASEB JOURNAL （ 影响因子：4.8； 五年影响因子：5.2 ）

ISSN： 0892-6638

年卷期： 2023 年 37 卷 8 期

页码：

收录情况： SCI

摘要： CRISPR-Cas9 is a versatile gene editing tool with a broad application of basic research and clinical therapeutics. However, the potential impact caused by off-target effects remains a critical bottleneck. The small Cas9 ortholog from Staphylococcus auricularis (SauriCas9) was identified, which recognizes a 5'-NNGG-3' protospacer adjacent motif (PAM), exhibiting high activity for genome editing. Recently, we also reported enhanced-fidelity Staphylococcus aureus Cas9 (efSaCas9), which harbors a single mutation N260D. Protein sequence alignment revealed that SauriCas9 has 62.4% sequence identity with SaCas9. Because SauriCas9 is more flexible in recognizing the target sequence with PAM of 5'-NNGG-3' than SaCas9 of 5'-NNGRRT-3' PAM, we sought to test whether key mutation(N260D) or adjacent residue mutation in efSaCas9 can be appliable to SauriCas9. With this concept, two engineered SauriCas9 variants (SauriCas9-HF1, harboring the N269D mutation; SauriCas9-HF2, harboring the D270N mutation) dramatically improved targeting specificity by targeted deep sequencing and GUIDE-seq. At certain sites, reduced off-target effects (approximately 61.6- and 111.9-fold improvements) of SauriCas9-HF2 compared with wild-type SauriCas9 were observed. Overall, two identified SauriCas9 variants (SauriCas9-HF1 and SauriCas9-HF2) expand the utility of the CRISPR toolkit for research and therapeutic applications.

分类号：

  • 相关文献
作者其他论文 更多>>

微信小程序