Fig. 3. Functional gametes were produced by the single blastomere induction of iPGCs.

Injection of 9GM or buc mRNA into a single blastomere at the animal pole (a) or margin (b) of a 128-cell embryo. GFP-UTRnanos3 was used to label iPGCs, and mCherryCAAX was used to label injected cells. c Time-lapse of iPGC migration to the genital ridges after single blastomere injection. d Single blastomere-induced iPGCs migrated to the genital ridge. e, f Genome editing was performed in one blastomere at 128-cell stage without iPGC induction. Eventually, these genome-edited cells migrated to other parts of the body besides the genital ridge. g Schematic representation of simultaneous genome editing and iPGC induction in a single blastomere at 128-cell stage. Embryos were injected with a low dose of dnd MO2 (20 µM) at 1-cell stage to eliminate the endogenous PGC, and the single blastomere induced iPGCs developed into genome-edited gametes. h Mutation efficiencies of gametes originated from single blastomere induced iPGCs. For each fish, a total of 10 embryos obtained from the hybridization of genome-edited sperm originated from single blastomere induced iPGCs and wild-type eggs were analyzed. The mutation efficiency of 5 F0 fish (#1–#5) gametes is shown for each gene. A representative example of three replicate is shown.