Rapid establishment of Oct4: EGFP transgenic zebrafish homozygote through gynogenesis for monitoring the pluripotency during induction of pluripotent stem cells

Abstract

Oct4 was one of the important markers of cellular pluripotency. In this study, by Tol2 transgenic technology, the recombinant plasmids composed of Oct4 promoter DNA and enhanced green fluorescent protein (EGFP) gene are microinjected into zebrafish 1-cell stage embryos, and obtain the EGFP positive (EGFP+) transgenic fish. To generation homozygous transgenic fish, the eggs of EGFP+ F0 were activated by ultraviolet inactivated sperm of crucian carp, and then inhibited their first cleavage. The homozygous Oct4-EGFP+ zebrafish are successfully obtained in F1 generation. After chemical reprogramming, the clones derived from caudal fin fibroblasts of homozygous Oct4-EGFP+ zebrafish are observed green fluorescence. These results suggest that gynogenetic technology provides new strategies for the rapid preparation of homozygous transgenic fish.