Harnessing the native type I-B CRISPR-Cas for genome editing in a polyploid archaeon  

作　　者：Feiyue Cheng Luyao Gong Dahe Zhao Haibo Yang Jian Zhou Ming Li Hua Xiang 

机构地区：[1]State Key Laboratory of Microbial Resources,Institute of Microbiology,Chinese Academy of Sciences,Beijing 100101,China [2]University of Chinese Academy of Sciences,Beijing 100049,China

出　　处：《Journal of Genetics and Genomics》2017年第11期541-548,共8页遗传学报（英文版）

基　　金：supported by the grants from the National Natural Science Foundation of China(No.31571283);the CASSAFEA International Partnership Program for Creative Research Teams

摘　　要：Research on CRISPR-Cas（clustered regularly interspaced short palindromic repeats-CRISPR associated protein） systems has led to the revolutionary CRISPR/Cas9 genome editing technique. However, for most archaea and half of bacteria, exploitation of their native CRISPR-Cas machineries may be more straightforward and convenient. In this study, we harnessed the native type I-B CRISPR-Cas system for precise genome editing in the polyploid haloarchaeon Haloarcula hispanica. After testing different designs, the editing tool was optimized to be a single plasmid that carries both the self-targeting miniCRISPR and a 600-800 bp donor. Significantly, chromosomal modifications, such as gene deletion, gene tagging or single nucleotide substitution, were precisely introduced into the vast majority of the transformants. Moreover, we showed that simultaneous editing of two genomic loci could also be readily achieved by one step. In summary, our data demonstrate that the haloarchaeal CRISPR-Cas system can be harnessed for genome editing in this polyploid archaeon, and highlight the convenience and efficiency of the native CRISPR-based genome editing strategy.Research on CRISPR-Cas（clustered regularly interspaced short palindromic repeats-CRISPR associated protein） systems has led to the revolutionary CRISPR/Cas9 genome editing technique. However, for most archaea and half of bacteria, exploitation of their native CRISPR-Cas machineries may be more straightforward and convenient. In this study, we harnessed the native type I-B CRISPR-Cas system for precise genome editing in the polyploid haloarchaeon Haloarcula hispanica. After testing different designs, the editing tool was optimized to be a single plasmid that carries both the self-targeting miniCRISPR and a 600-800 bp donor. Significantly, chromosomal modifications, such as gene deletion, gene tagging or single nucleotide substitution, were precisely introduced into the vast majority of the transformants. Moreover, we showed that simultaneous editing of two genomic loci could also be readily achieved by one step. In summary, our data demonstrate that the haloarchaeal CRISPR-Cas system can be harnessed for genome editing in this polyploid archaeon, and highlight the convenience and efficiency of the native CRISPR-based genome editing strategy.

关 键 词：Haloarcula hispanica CRISPR-Cas Genome editing Polyploid 

分 类 号：Q78[生物学—分子生物学]