【摘要】 漠斑牙鲆(Paralichthys lethostigma)是中国近年新开发的重要海水养殖鱼类,本研究以漠斑牙鲆囊胚期胚胎为材料,探索了鱼类胚胎细胞分离和培养的条件,建立了漠斑牙鲆胚胎细胞培养技术。建立的漠斑牙鲆胚胎细胞系(SFEC)至今已经培养了240多天,传至80多代。SFEC的完全培养基采用添加抗生素、胎牛血清(FBS)、花鲈血清(SPS)、成纤维生长因子(bFGF)的DMEM。SFEC形态小而呈圆形、多边形,在培养基中生长迅速。本实验检测了温度、胎牛血清浓度、成纤维生长因子对SFEC生长的影响。在24~30℃之间SFEC生长良好,但当温度低于18℃时细胞生长速度明显减慢。在含15%FBS的培养基中,SFEC生长速度明显比在含7.5%FBS的培养基中快,在培养基中添加bFGF可以使细胞生长速度显著提高。SFEC的二倍体核型为2n=6st+42t。将GFP报告基因通过脂质体介导的方法转入SFEC中并成功地获得了表达,转化率为20%。更多还原

【Abstract】 A continuous cell line,SFEC(southern flounder embryonic cell line),was established from embryos at gastrula stage of a marine cultured fish,southern flounder(Paralichthys lethostigma) and was cultured for more than 240d with more than 80 generation.The SFEC cells were cultured in DMEM medium supplemented with antibiotics,fetal bovine serum(FBS),sea perch serum(SPS),basic fibroblast growth factor(bFGF).The cells were small and round in shape and grew actively and stably in medium.Effects of temperature,FBS and bFGF concentration on the growth of SFEC cells were evaluated.The cells grew well in the temperature range of 24-30℃,but had a reduced growth rate at temperature below 18℃.The growth of SFEC cells in medium containing 15% FBS was higher than that in medium containing 7.5% FBS.Addition of bFGF to the medium significantly increased growth rate of SFEC cells.Chromosome analysis revealed that SFEC cells have a normal diploid karyotype with 2n=6st+42t.GFP reporter gene was successfully transferred into SFEC cells and expressed.更多还原