ABSTRACT.- Microinjection of CRISPR/Cas9 requires the availability of zygotes that implies animal breeding, superovulation schemes, and embryo collection. Vitrification of zygotes may allow having ready-to-use embryos and to temporally dissociate the workload of embryo production from microinjection. In this study, fresh (F group) or vitrified (V group) zygotes were microinjected with CRISPR/Cas9 system to test the hypothesis that vitrified zygotes could be a suitable source of embryos for microinjection. In Experiment 1 (in vitro evaluation), B6D2F1/J zygotes were microinjected and cultured until blastocyst stage. © 2024 Schlapp et al.