CRISPR/Cas9 Gene Editing of Turkey Cells Using Adenoviral Delivery Running Head: RESEARCH NOTE

Poultry, including birds such as chicken, duck, turkey, geese, and other game birds is the most widely consumed animal protein globally, with per capita poultry consumption steadily rising in the United States over the past several decades. To meet the demand for poultry, many avenues of improving poultry production have been explored including genetic modification solutions. While genetic modification in many poultry species has been well explored, gene editing in turkey has no research into its application. This study investigated the feasibility of using CRISPR/Cas9 mediated gene editing using adenoviral delivery in vitro, a method that has been proven successful for other species of poultry but never turkey. Three guide RNAs (gRNAs) were designed, targeting the leptin receptor (LEPR) gene matching the gene sequences of both chicken and turkey. Following the assessment of each gRNA's editing efficiency in turkey embryonic fibroblast (TEF) cells and commercially available chicken DF-1 cells via liposome-mediated transfection of the CRISPR vector, the most effective gRNA was chosen for recombinant adenovirus type 5 production. From there, both DF-1 and TEF cells were transduced using the adenovirus and the genome editing efficiency was assessed. The results show that turkey cells can be genetically modified in vitro with indel frequencies comparable to those observed in chicken cells, using both CRISPR plasmids and adenoviral vectors. These findings can lay the groundwork for future experiments and adaptation to in vivo modification of turkey in the future.