Oligonucleotide sequences.

<div><p>Fibroblast growth factor receptor 4 (FGFR4) has a role in many biological processes, including lipid metabolism, tissue repair, and vertebrate development. In recent years, FGFR4 overexpression and activating mutations have been associated with numerous adult and pediatric cancers. As such, FGFR4 presents an opportunity for therapeutic targeting which is being pursued in clinical trials. To understand the role of FGFR4 signaling in disease and development, we generated and characterized three alleles of <i>fgfr4</i> knockout zebrafish strains using CRISPR/Cas9. To generate <i>fgfr4</i> knockout crispants, we injected single-cell wildtype zebrafish embryos with <i>fgfr4</i> targeting guide RNA and Cas9 proteins, identified adult founders, and outcrossed to wildtype zebrafish to create an F1 generation. The generated mutations introduce a stop codon within the second Ig-like domain of Fgfr4, resulting in a truncated 215, 223, or 228 amino acid Fgfr4 protein compared to 922 amino acids in the full-length protein. All mutant strains exhibited significantly decreased <i>fgfr4</i> mRNA expression during development, providing evidence for successful knockout of <i>fgfr4</i> in mutant zebrafish. We found that, consistent with other <i>Fgfr4</i> knockout animal models, the <i>fgfr4</i> mutant fish developed normally; however, homozygous <i>fgfr4</i> mutant zebrafish were significantly smaller than wildtype fish at three months post fertilization. These <i>fgfr4</i> knockout zebrafish lines are a valuable tool to study the role of FGFR4 in vertebrate development and its viability as a potential therapeutic target in pediatric and adult cancers, as well as other diseases.</p></div>