Efficient CRISPR/Cas9 genome editing in a salmonid fish cell line using a lentivirus delivery system.

Autor: Gratacap RL; The Roslin Institute, University of Edinburgh, Easter Bush campus, Midlothian, UK. Remi.Gratacap@ed.ac.uk., Regan T; The Roslin Institute, University of Edinburgh, Easter Bush campus, Midlothian, UK., Dehler CE; Institute of Biological and Environmental Sciences, University of Aberdeen, Aberdeen, UK., Martin SAM; Institute of Biological and Environmental Sciences, University of Aberdeen, Aberdeen, UK., Boudinot P; Virologie et Immunologie Moleculaires, Institut National de Recherche Agronomique (INRA), Universite Paris-Saclay, Jouy-en-Josas, France., Collet B; Virologie et Immunologie Moleculaires, Institut National de Recherche Agronomique (INRA), Universite Paris-Saclay, Jouy-en-Josas, France., Houston RD; The Roslin Institute, University of Edinburgh, Easter Bush campus, Midlothian, UK. Ross.Houston@roslin.ed.ac.uk.
Jazyk: angličtina
Zdroj: BMC biotechnology [BMC Biotechnol] 2020 Jun 23; Vol. 20 (1), pp. 35. Date of Electronic Publication: 2020 Jun 23.
DOI: 10.1186/s12896-020-00626-x
Abstrakt: Background: Genome editing is transforming bioscience research, but its application to non-model organisms, such as farmed animal species, requires optimisation. Salmonids are the most important aquaculture species by value, and improving genetic resistance to infectious disease is a major goal. However, use of genome editing to evaluate putative disease resistance genes in cell lines, and the use of genome-wide CRISPR screens is currently limited by a lack of available tools and techniques.
Results: In the current study, we developed an optimised protocol using lentivirus transduction for efficient integration of constructs into the genome of a Chinook salmon (Oncorhynchus tshwaytcha) cell line (CHSE-214). As proof-of-principle, two target genes were edited with high efficiency in an EGFP-Cas9 stable CHSE cell line; specifically, the exogenous, integrated EGFP and the endogenous RIG-I locus. Finally, the effective use of antibiotic selection to enrich the successfully edited targeted population was demonstrated.
Conclusions: The optimised lentiviral-mediated CRISPR method reported here increases possibilities for efficient genome editing in salmonid cells, in particular for future applications of genome-wide CRISPR screens for disease resistance.
Databáze: MEDLINE
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