Development and Characterization of Embryonic Stemlike Cell Culture from Carassius auratas (Linnaeus, 1758)

Abstract

Embryonic stem cells are pluripotent cells, capable of differentiating into various cell types. In the present study, feeder cell-free culture of midblastula embryos of the Goldfish (Carassius auratus) was carried out. Embryonic stem cells were derived from mid-blastula stage embryos using pronase to release the inner cell mass (ICM). The cells were cultured in Leibovitz-15 medium on gelatin-coated cell culture well plates supplemented with 15% fetal bovine serum, 1% goldfish serum, basic fibroblast growth factor and leukaemia inhibitory factors (LIF) and were incubated at 28°C. The ES-like cells were characterized by their unique round and polygonal morphology, elevated activity of alkaline phosphatase, spherical three-dimensional embryoid body formation, chromosomal diploid number (2n=50). The expression of Oct4, a member Oct family of POU transcription factors, which play a key role in regulating stem cell pluripotency and differentiation, was used for characterization of undifferentiated pluripotent ES cells. The expression of Oct4 in embryonic stem-like cell (2nd passage) and gonads of goldfish was studied by RT- PCR. Embryonic stem-like cells culture from the mid blastula embryo of C. auratus would be a useful tool for cellular development and differentiation studies.