Development and characterization of embryonic stem-like cell culture from Carassius auratas (Linnaeus, 1758).

Abstract

Embryonic stem cells are pluripotent cells, capableof differentiating into various cell types. In thepresent study, feeder cell-free culture of mid-blastula embryos of the Goldfish (Carassius auratus)was carried out. Embryonic stem cells were derivedfrom mid-blastula stage embryos using pronase torelease the inner cell mass (ICM). The cells werecultured in Leibovitz-15 medium on gelatin-coatedcell culture well plates supplemented with 15% fetalbovine serum, 1% goldfish serum, basic fibroblastgrowth factor and leukaemia inhibitory factors (LIF)and were incubated at 28°C. The ES-like cells werecharacterized by their unique round and polygonalmorphology, elevated activity of alkaline phos-phatase, spherical three-dimensional embryoid bodyformation, chromosomal diploid number (2n=50).The expression of Oct4, a member Oct family ofPOU transcription factors, which play a key role inregulating stem cell pluripotency and differentia-tion, was used for characterization of undifferenti-ated pluripotent ES cells. The expression of Oct4 inembryonic stem-like cell (2nd passage) and gonadsof goldfish was studied by RT- PCR. Embryonicstem-like cells culture from the mid blastula embryoof C. auratus would be a useful tool for cellulardevelopment and differentiation studies.