Genetic Engineering in Model Livestock Species

Background: Genetically engineered farm animals are attractivealternative mammalian models to rodents for the study ofdevelopmental, genetic, reproductive and disease-related biologicalquestions. However until recently, the molecular methodologiesfor genetic engineering in these species were notestablished. The development, refinement and optimization ofmolecular methods for precise genetic modifications are necessaryto fully exploit the potential of these species for translationalresearch.Material, Methods and Results: DNA-Transposons, Recombinasesand Crispr/Cas۹ designer nucleases have been employedfor direct injection in murine, porcine and bovine zygotesto achieve precise genetic modifications.Gain-of-function and loss-of-function modifications have beenachieved, and a simplified injection technique has been established.Importantly, the cytoplasmic plasmid injection (CPI) isalso suitable for opaque zygotes from pigs or cows.Conclusions: Although the transgenic toolbox for large animalsis currently equipped with powerful methodologies thereare many aspects to improve in the associated reproductivetechnologies. Low success rates of animal cloning (SCNT) andzygote microinjection, two of the most commonly used method,still represent a bottleneck. The ground-breaking feature of theCrispr/Cas۹ system is that it brought the possibility of purposelydirecting the genomic modification to a specific chromosomallocus. It is anticipated that new generation transgenic tools inconcert with updated genomic data will facilitate the production of large animal models for translational medicine. These largeanimal models will be instrumental for understanding diseasepathogenesis and development of better therapeutic approachesof severe human pathologic conditions.