内蒙古绒山羊原始生殖细胞(PGCs)的培养与鉴定

Culture and Identification of Primordial Germ Cells from Inner Mongolia Cashmere Goat

本试验从内蒙古白绒山羊胎儿的生殖嵴中分离出原始生殖细胞体外培养并进行了鉴定。为建立内蒙古绒山羊原始生殖细胞(pri mordial germcells,PGCs)的培养体系,试验对比了A、B、C3组不同的培养条件对内蒙古绒山羊原始生殖细胞传代数的影响。RT-PCR鉴定结果表明,β-actin、hTERT、GAPDH、Nanog、Oct3/4为阳性、AKP染色为阳性;免疫荧光检测胚胎干细胞特异标志物Oct3/4、SSEA-3、SSEA-4呈阳性。未添加任何因子的A培养体系仅能传1代,添加LIF及Insulin的B培养体系可以传至4代,添加LIF、Insulin及优质胎牛血清的C组培养体系可以传至9代。

In this study,we cultured and identified of primordial germ cells（PGCs） which were isolated from genital ridge of the Inner Mongolia cashmere goat fetus.Three groups（A,B,C） culture medium were detected establish culture system for culture of primordial germ cells from Inner Mongolia cashmere goat.After identified by RT-PCR,AKP staining and immunofluorescence,the results showed that the RT-PCR detection of β-actin,hTERT,GAPDH,Nanog and Oct3/4,AKP identified was positive,immunofluorescence detection of Oct3/4,SSEA-3,SSEA-4 were positive.Group A culture medium without any factors could only be passaged 1 generation,group B culture medium supplemented with LIF and insulin could be passaged 4 generations,group C culture medium supplemented with LIF,insulin and high-quality fetal bovine serum culture system could be passaged 9 generations.