Efficient Genome Editing in Multiple Salmonid Cell Lines Using Ribonucleoprotein Complexes
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ORIGINAL ARTICLE
Efficient Genome Editing in Multiple Salmonid Cell Lines Using Ribonucleoprotein Complexes Remi L. Gratacap 1
&
Ye Hwa Jin 1
&
Marina Mantsopoulou 1 & Ross D. Houston 1
Received: 3 April 2020 / Accepted: 31 August 2020 / Published online: 18 September 2020 # The Author(s) 2020
Abstract Infectious and parasitic diseases have major negative economic and animal welfare impacts on aquaculture of salmonid species. Improved knowledge of the functional basis of host response and genetic resistance to these diseases is key to developing preventative and treatment options. Cell lines provide valuable models to study infectious diseases in salmonids, and genome editing using CRISPR/Cas systems provides an exciting avenue to evaluate the function of specific genes in those systems. While CRISPR/Cas editing has been successfully performed in a Chinook salmon cell line (CHSE-214), there are no reports to date of editing of cell lines derived from the most commercially relevant salmonid species Atlantic salmon and rainbow trout, which are difficult to transduce and therefore edit using lentivirus-mediated methods. In the current study, a method of genome editing of salmonid cell lines using ribonucleoprotein (RNP) complexes was optimised and tested in the most commonly used salmonid fish cell lines: Atlantic salmon (SHK-1 and ASK cell lines), rainbow trout (RTG-2) and Chinook salmon (CHSE-214). Electroporation of RNP based on either Cas9 or Cas12a was efficient at targeted editing of all the tested lines (typically > 90% cells edited), and the choice of enzyme expands the number of potential target sites for editing within the genomes of these species. These optimised protocols will facilitate functional genetic studies in salmonid cell lines, which are widely used as model systems for infectious diseases in aquaculture. Keywords CRISPR . Salmonid . Cell line . Ribonucleoprotein . Genome editing . Disease resistance
Introduction Salmonid fish are amongst the highest value aquaculture species globally, together worth in excess of $22Bn in 2017 (FAO 2019). However, infectious disease outbreaks are a continuous threat to sustainable production and future expansion to meet global demands for these fish. Therefore, development of vaccines and therapeutics is an important goal, and selective breeding for improved host resistance has major potential to help tackle several diseases (Yáñez et al. 2014). Genomic selection has also been applied to enhance the rate of genetic Remi L. Gratacap and Ye Hwa Jin contributed equally to this work. Electronic supplementary material The online version of this article (https://doi.org/10.1007/s10126-020-09995-y) contains supplementary material, which is available to authorized users. * Ross D. Houston [email protected] 1
The Roslin Institute and Royal (Dick) School of Veterinary Studies, University of Edinburgh, Midlothian EH25 9RG, UK
gain for disease resistance traits in breeding programmes (Houston 2017; Zenger et al. 2019; Houston et al. 2020), and genome editi
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