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Genome Res. 2020 Jan;30(1):107-117. doi: 10.1101/gr.255414.119. Epub 2020 Jan 3.

Versatile and robust genome editing with Streptococcus thermophilus CRISPR1-Cas9.

Author information

1
Centre Hospitalier Universitaire de Québec Research Center-Université Laval, Québec, Québec G1V 4G2, Canada.
2
Centre de recherche de l'Institut universitaire de cardiologie et de pneumologie de Québec (CRIUCPQ)-Université Laval, Québec, Québec G1V 4G5, Canada.
3
Service de Génétique médicale, Département de Pédiatrie, Centre Hospitalier Universitaire de Sherbrooke (CHUS), et CRCHUS, Sherbrooke, Québec J1H 5N4, Canada.
4
Université Laval Cancer Research Centre, Québec, Québec G1V 0A6, Canada.
5
Département de biochimie, de microbiologie, et de bio-informatique, Faculté des sciences et de génie, Université Laval, Québec, Québec G1V 0A6, Canada.
6
Groupe de recherche en écologie buccale, Faculté de médecine dentaire, Université Laval, Québec, Québec G1V 0A6, Canada.
7
Félix d'Hérelle Reference Center for Bacterial Viruses, Faculté de médecine dentaire, Université Laval, Québec, Québec G1V 0A6, Canada.
8
Architecture et Fonction des Macromolécules Biologiques, Centre National de la Recherche Scientifique (CNRS), Campus de Luminy, 13288 Marseille Cedex 09, France.
9
Architecture et Fonction des Macromolécules Biologiques, Aix-Marseille Université, Campus de Luminy, 13288 Marseille Cedex 09, France.

Abstract

Targeting definite genomic locations using CRISPR-Cas systems requires a set of enzymes with unique protospacer adjacent motif (PAM) compatibilities. To expand this repertoire, we engineered nucleases, cytosine base editors, and adenine base editors from the archetypal Streptococcus thermophilus CRISPR1-Cas9 (St1Cas9) system. We found that St1Cas9 strain variants enable targeting to five distinct A-rich PAMs and provide a structural basis for their specificities. The small size of this ortholog enables expression of the holoenzyme from a single adeno-associated viral vector for in vivo editing applications. Delivery of St1Cas9 to the neonatal liver efficiently rewired metabolic pathways, leading to phenotypic rescue in a mouse model of hereditary tyrosinemia. These robust enzymes expand and complement current editing platforms available for tailoring mammalian genomes.

PMID:
31900288
DOI:
10.1101/gr.255414.119

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