CRISPR/dCas9 Knock-down and CRISPR/Cas9 Knock-out of lxr in Atlantic salmon SHK-1 cells
Abstract
Understanding the genetics of Atlantic salmon (Salmo salar) is crucial for gaining insight into this commercially important species. The sustainable development of aquaculture requires increasing levels of plant-based ingredients in fish diets, which lack cholesterol and have less essential fatty acids that are important for consumers. The lxr gene, a member of the nuclear hormone receptor family, has shown to be a key regulator of lipid metabolism in vertebrates, also in Atlantic salmon (Minghetti et al., 2011). Despite its recognized importance, the precise mechanisms of action and downstream targets of lxr in Atlantic salmon remain incompletely understood. To address this deficiency, we have investigated the role of lxr in SHK-1 cells in Atlantic salmon using CRISPR/Cas9 gene editing techniques to knock-out the gene and analyze the resulting changes in mRNA levels. Furthermore, we wanted to study the potential application of CRISPR/dCas9, a modified version of CRISPR/Cas9 that can be utilized for transcriptional regulation in Atlantic salmon. To the best of our knowledge, the application of knock-down by CRISPR/dCas9 in Atlantic salmon has not been reported previously. The sample number in this thesis was too low to conclude if the CRISPR/dCas9 system can be functional in Atlantic salmon, but the results showed indications of a successful lxr downregulation. Thus, more tests with a larger number of samples are needed to confirm these findings. The knock-out of the lxr gene resulted in the upregulation of 25 genes and the downregulation of 19 genes.