title: Harnessing Type I and Type III CRISPR-Cas systems for genome editing.
abstract: CRISPR-Cas (clustered regularly interspaced short palindromic repeats-CRISPR-associated) systems are widespread in archaea and bacteria, and research on their molecular mechanisms has led to the development of genome-editing techniques based on a few Type II systems. However, there has not been any report on harnessing a Type I or Type III system for genome editing. Here, a method was developed to repurpose both CRISPR-Cas systems for genetic manipulation in Sulfolobus islandicus, a thermophilic archaeon. A novel type of genome-editing plasmid (pGE) was constructed, carrying an artificial mini-CRISPR array and a donor DNA containing a non-target sequence. Transformation of a pGE plasmid would yield two alternative fates to transformed cells: wild-type cells are to be targeted for chromosomal DNA degradation, leading to cell death, whereas those carrying the mutant gene would survive the cell killing and selectively retained as transformants. Using this strategy, different types of mutation were generated, including deletion, insertion and point mutations. We envision this method is readily applicable to different bacteria and archaea that carry an active CRISPR-Cas system of DNA interference provided the protospacer adjacent motif (PAM) of an uncharacterized PAM-dependent CRISPR-Cas system can be predicted by bioinformatic analysis.
authors.affiliation: State Key Laboratory of Agricultural Microbiology, College of Life Science and Technology, Huazhong Agricultural University, Wuhan, Hubei 430070, China.
authors.forename: Yingjun
authors.initials: Y
authors.lastname: Li
authors.affiliation: State Key Laboratory of Agricultural Microbiology, College of Life Science and Technology, Huazhong Agricultural University, Wuhan, Hubei 430070, China.
authors.forename: Saifu
authors.initials: S
authors.lastname: Pan
authors.affiliation: State Key Laboratory of Agricultural Microbiology, College of Life Science and Technology, Huazhong Agricultural University, Wuhan, Hubei 430070, China.
authors.forename: Yan
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: State Key Laboratory of Agricultural Microbiology, College of Life Science and Technology, Huazhong Agricultural University, Wuhan, Hubei 430070, China.
authors.forename: Min
authors.initials: M
authors.lastname: Ren
authors.affiliation: State Key Laboratory of Agricultural Microbiology, College of Life Science and Technology, Huazhong Agricultural University, Wuhan, Hubei 430070, China.
authors.forename: Mingxia
authors.initials: M
authors.lastname: Feng
authors.affiliation: State Key Laboratory of Agricultural Microbiology, College of Life Science and Technology, Huazhong Agricultural University, Wuhan, Hubei 430070, China.
authors.forename: Nan
authors.initials: N
authors.lastname: Peng
authors.affiliation: Key Laboratory of Quality and Safety Risk Assessment for Aquatic Products on Storage and Preservation (Shanghai), China Ministry of Agriculture; College of Food Science and Technology, Shanghai Ocean University, 999 Hu Cheng Huan Road, Shanghai 201306, China.
authors.forename: Lanming
authors.initials: L
authors.lastname: Chen
authors.affiliation: State Key Laboratory of Agricultural Microbiology, College of Life Science and Technology, Huazhong Agricultural University, Wuhan, Hubei 430070, China fa-lyx@163.com.
authors.forename: Yun Xiang
authors.initials: YX
authors.lastname: Liang
authors.affiliation: State Key Laboratory of Agricultural Microbiology, College of Life Science and Technology, Huazhong Agricultural University, Wuhan, Hubei 430070, China Archaea Centre, Department of Biology, University of Copenhagen, Ole Maaløes Vej 5, DK2200 Copenhagen N, Denmark qunxin@bio.ku.dk.
authors.forename: Qunxin
authors.initials: Q
authors.lastname: She
mesh_terms: D064113:CRISPR-Cas Systems / Q000235:genetics*; D004247:DNA / Q000235:genetics; D020745:Genome, Archaeal; D010957:Plasmids / Q000235:genetics; D017393:RNA Editing / Q000235:genetics*; D016944:Sulfolobus / Q000235:genetics*
chemical_list: D004247:DNA