Establishment and characterization of an ovarian cell line from half-smooth tongue sole Cynoglossus semilaevis

A new ovarian cell line, CSO, was established from half-smooth tongue sole Cynoglossus semilaevis. Primary culture of CSO cells was initiated from digestion of ovarian tissues pieces by trypsin solution and cultured at 24° C in Dulbecco's modified Eagle's medium-F12 medium (DMEM-F12, 1:1) (pH 7·0), supplemented with 20% foetal bovine serum, basic fibroblast growth factor (bFGF), epidermal growth factor (EGF), insulin-like growth factor-I (IGF-I) and human chorionic gonadotropin (HCG). The cultured CSO cells, fibroblastic in morphology, proliferated to 100% confluency 3 days later and had been subcultured to passage 80. Chromosome analyses indicated that the CSO cells exhibited chromosomal aneuploidy with a modal chromosome number of 42 that displayed the normal diploid karyotype of C. semilaevis [2n = 42 t, fundamental number (NF ) = 42]. Reverse transcription polymerase chain reaction revealed that CSO cells could express ovarian somatic cell functional genes p450armo, foxl2 and sox9a but not ovary germ cell marker gene vasa and male-specific gene dmrt1. Transfection experiment demonstrated that CSO cells transfected with pEGFP-N3 plasmid could express green fluorescence protein (GFP) with higher transfection efficiency. The CSO cell line might serve as a valuable tool for studies on the mechanism of sex determination and oogenesis of ovary in flatfish.