半滑舌鳎肝脏细胞系的建立与鉴定

Development and characterization of a liver cell line from half-smooth tongue-sole(Cynoglossus semilaevis)

以半滑舌鳎肝脏组织为材料,探索了组织细胞分离培养条件,建立了半滑舌鳎组织细胞培养技术及半滑舌鳎肝脏细胞系(HTLC)。该细胞系的培养基是添加了抗生素、胎牛血清(FBS)、花鲈血清(SPS)、成纤维生长因子(bFGF)的 MEM。HTLC 形态呈纤维状,在培养基中生长迅速,经200多天的培养,成功传代30多代。检测了温度、FBS 浓度、bFGF对其生长的影响,结果表明,在24～30℃生长良好,但当温度低于12℃时细胞生长速度明显减慢;在一定浓度范围内,其生长速度随血清浓度的升高而增快,血清浓度过高或过低对其生长不利;在培养基中添加 bFGF 可以使细胞生长速度显著提高;其二倍体核型为2n=21t。将绿色荧光蛋白(GFP)报告基因通过脂质体介导的方法转入 HTLC 中并成功地获得了表达,转化率为20%左右。该细胞系的建立为研究鱼类病毒学、免疫学、遗传学、功能基因组学提供了很好的实验材料。

A liver cell line from half-smooth tongue-sole （ Cynoglossus semilaevis ）, named HTLC, was established and was cultured for more than 200 days with more than 30 passages. The HTLC cells were cultured in the MEM medium supplemented with antibiotics, fetal bovine serum （FBS）, sea perch serum （SPS）, and basic fibroblast growth factor （bFGF）. The cells were fibriform and stably grew in culture. The effects of temperature, FBS concentration and bFGF on the growth of HTLE cells were examined. The cells grew well in the temperature range of 24-30℃, but had a reduced growth rate at the temperature below 12℃. The growth rate of HTLC cells in the medium containing 20% FBS was higher than that in the medium containing 15% and 10% FBS. Addition of bFGF to the medium significantly increased the growth rate of HTLC cells. Chromosome analysis revealed that HTLC cells had a normal diploid karyotype with 2n = 21t. Green fluorescent protein （GFP） reporter genes were transferred into HTLC cells and were successfully expressed.