FIGURE SUMMARY
Title

Generation of germ-line chimera zebrafish using primordial germ cells isolated from cultured blastomeres and cryopreserved embryoids

Authors
Kawakami, Y., Goto-Kazeto, R., Saito, T., Fujimoto, T., Higaki, S., Takahashi, Y., Arai, K., and Yamaha, E.
Source
Full text @ Int. J. Dev. Biol.
Fig. 1

Visualized green fluorescent protein (GFP)-positive cells obtained using the dissociated blastomere (DB) and cultured embryoid (CE) methods. (A) Isolation of blastodiscs at the late-blastula stage from embryos injected with GFP-nos1-3′UTR strand-capped mRNA. Large embryos were blastulae before operation, whereas small ones were isolated blastodiscs. (B) DBs cultured for 1 day in a dissociated condition in sodium citrate solution. (C) Embryoid from an isolated blastodisc cultured for 1 day in Ringer?s solution. Arrows indicate GFPpositive cells (primordial germ cells). Bars, 500 μm.

Fig. 2

Migration and proliferation of donor primordial germ cells (PGCs) derived from dissociated blastomeres cultured for 1 day at 26ºC in the same germ-line chimera. Green fluorescent protein-positive cells indicate donor PGCs and DsRed-positive cells indicate host PGCs. (A) Host at 1 day post fertilization (dpf). (B) 2 dpf. (C) 3 dpf. (D) 11 dpf. Bars, 500 μm.

Fig. 3

Migration and proliferation of donor primordial germ cells (PGCs) derived from cryopreserved/ thawed embryoids cultured for 1 day at 22ºC in the same germ-line chimera. Arrows indicate green fluorescent protein-labeled PGCs or germ cells. (A) Host at 1 day post-fertilization (dpf). (B) 2 dpf. (C) 3 dpf. (D) 5 dpf. (E) 10 dpf. (F) 13 dpf. (G) 18 dpf. Bars, 500 μm.

Fig. 4

Zebrafish used in this study and melanophore pigmentation of control and F1 offspring from a cross between normal golden-type females and chimeric male fish. (A) Control offspring of a wild-type zebrafish at 2 days postfertilization (dpf). (B) Control offspring of a goldentype zebrafish at 2 dpf. (C) Control offspring of a cross between a golden-type female and wild-type male at 2 dpf. Phenotype is wild-type. (D) Offspring at 2 dpf of a cross between a golden-type female and a golden-type male chimera, in which a wild-type primordial germ cell (PGC) from an embryoid cultured for 2 days at 22°C was transplanted into a golden-type blastula. Phenotype is wild-type. (E) Offspring at 2 dpf of a cross between a golden-type female and a golden-type male chimera, in which a wild-type PGC from a cryopreserved/thawed embryoid cultured for 1 day at 22°C was transplanted into a golden-type blastula. Phenotype is wild-type. (F) Control offspring at 30 dpf of a cross between a golden-type female and a wild-type male. (G) Control offspring at 30 dpf of a cross between a golden-type female and a golden-type male. (H) Control offspring at 30 dpf of a cross between a wild-type male and a golden-type female. (I) Offspring at 30 dpf of a cross between a golden-type female and a goldentype male chimera transplanted with a wild-type PGC from a cryopreserved/thawed embryoid cultured for 1 day at 22°C. Bars in (A,B,C,D,E), 500 μm; Bars in (F,G,H,I), 2 mm.

Acknowledgments
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