期刊
BMC BIOTECHNOLOGY
卷 20, 期 1, 页码 -出版社
BMC
DOI: 10.1186/s12896-020-00626-x
关键词
CRISPR; Lentivirus; Gene editing; CHSE; Salmon; Disease resistance
资金
- Biotechnology and Biological Sciences Research Council [BB/R008612/1, BB/S004343/1, BB/R008973/1]
- Institute Strategic Programme Grants [BBS/E/D/20002172, BBS/E/D/30002275, BBS/E/D/10002070]
- BBSRC [BB/P017223/1, BB/R008973/1, BB/S004343/1, BBS/E/D/10002071, BB/R008612/1] Funding Source: UKRI
- NERC [NE/P010946/1] Funding Source: UKRI
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.
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