Derivation and Characterization of a New Embryonic Cell Line from the Olive Flounder Paralichthys olivaceus

Abstract

A new embryonic cell line that consisted predominantly of fibroblast cells has been established from embryos at Kupffer’s vesicle (KV) stage of the olive flounder Paralichthys olivaceus, designated as the PoEKC. PoEKC cells have been subcultured for 61 passages over a period of 18 months. The cells were cultured in DMEM/F-12 medium supplemented with antibiotics, FBS, and growth factors at temperature of 25°C. The growth curve of the PoEKC cells comprised an exponential growth phase and a long plateau phase. Chromosome analysis revealed that the cells possessed the normal flounder diploid karyotype of 2n = 48t. The origin of the cell line was confirmed by testing the partial sequences of cytochrome oxidase c subunit I (COI) gene of mtDNA. The pluripotency markers, genes OCT4-1, SOX2, KLF4, and NANOG showed positive signals in the PoEKC cells, and the cells also presented high ALP activity. According to the above results, the PoEKC cells might be pluripotency, though the pluripotency needs further confirmation. The cells were also successfully transfected with GFP reporter gene suggesting that it could be utilized for gene function study in the flounder.