Fig. 4

Schematic showing the number of steps required to produce live, homozygous, non-mosaic, GnEd livestock (maroon calf) through either somatic cell nuclear transfer (SCNT) cloning (tan arrows) or zygote microinjection (light purple arrows). Both methods include gamete collection and maturation, introduction of the gene-editing (GnEd) reagents, and transfer of embryos into synchronized recipients (surrogate dams). For the SCNT cloning approach (tan arrows) GnEd reagents are introduced into a somatic cell line and then SCNT cloning is used to produce embryos for transfer. The GnEd cell line can be screened before cloning to ensure production of a homozygous, non-mosaic animal. For the zygote microinjection approach (light purple arrows) GnEd reagents are introduced directly into a zygote via cytoplasmic injection or electroporation. GnEd of zygotes can result in mosaic offspring, which requires subsequent breeding to produce first heterozygous and ultimately homozygous GnEd offspring. Therefore, gene editing of zygotes may require more steps to produce a homozygous, non-mosaic, GnEd animal, as indicated by the increased number of light purple arrows (7) compared to the number of tan arrows (3). Reproduced from (Bishop and Van Eenennaam 2020) under a CC-BY license