An engineered Cas12i nuclease that is an efficient genome editing tool in animals and plants  

作　　者：Zhiqiang Duan Yafeng Liang Jialei Sun Hongjin Zheng Tong Lin Pengyu Luo Mengge Wang Ruiheng Liu Ying Chen Shuhua Guo Nannan Jia Hongtao Xie Meili Zhou Minghui Xia Kaijun Zhao Shuhui Wang Na Liu Yongling Jia Wei Si Qitong Chen Yechun Hong Ruilin Tian Jian-Kang Zhu 

机构地区：[1]Institute of Advanced Biotechnology and School of Medicine,Southern University of Science and Technology,Shenzhen 518055,China [2]Bellagen Biotechnology,Jinan 250000,China [3]Department of Medical Neuroscience,School of Medicine,Key University Laboratory of Metabolism and Health of Guangdong,Southern University of Science and Technology,Shenzhen,Guangdong Province 518055,China

出　　处：《The Innovation》2024年第2期61-69,共9页创新（英文）

基　　金：This work was supported by Shandong BellaGen Biotechnology,and by a grant from the National Natural Science Foundation of China(32188102 to J.-K.Z.).

摘　　要：The type V-I CRISPR-Cas system is becoming increasingly more attractive for genome editing.However,natural nucleases of this system often exhibit low efficiency,limiting their application.Here,we used structure-guided rational design and protein engineering to optimize an uncharacterized Cas12i nuclease,Cas12i3.As a result,we developed Cas-SF01,a Cas12i3 variant that exhibits significantly improved gene editing activity in mammalian cells.Cas-SF01 shows comparable or superior editing performance compared to SpCas9 and other Cas12 nucleases.Compared to natural Cas12i3,Cas-SF01 has an expanded PAM range and effectively recognizes NTTN and noncanonical NATN and TTVN PAMs.In addition,we identified an amino acid substitution,D876R,that markedly reduced the off-target effect while maintaining high on-target activity,leading to the development of CasSF01^(HiFi)(high-fidelity Cas-SF01).Finally,we show that Cas-SF01 has high gene editing activities in mice and plants.Our results suggest that CasSF01 can serve as a robust gene editing platform with high efficiency and specificity for genome editing applications in various organisms.

关 键 词：system ATTRACTIVE rational 

分 类 号：Q814[生物学—生物工程] Q78