第28期鸡胚原始生殖细胞的冷冻保存与体外培养研究

利用Ficoll密度梯度离心,提取鸡胚第28期(孵化132h)性腺中的原始生殖细胞(PGCs),用不同的冷冻保护液,对PGCs采用分离后直接冷冻保存和体外培养24h后冷冻保存2种方法,以筛选合适的冷冻保护体系。结果发现,分离提纯后直接进行冷冻保存的PGCs,存活率最高为(87.07±1.29)%;体外培养24h后进行冷冻的PGCs,存活率最高为(44.08±1.19)%。对复苏后的PGCs进行体外培养结果发现,分离提纯后直接进行冷冻的PGCs,在体外培养过程中具有形成细胞克隆的能力,且可传代和进行体外分化;而体外培养24h后进行冷冻的PGCs,复苏后在体外培养过程中不能形成细胞克隆,且在体外培养40h左右死亡。

Cryopreservation and culture of chicken primordial germ cells at stage 28

In order to investigate the possibility of cryopreservation and differentiation of the chicken primordial germ cells (PGCs) at stage 28,the PGCs from the gonada by Ficoll density-gradient centrifugation were isolated,and cryopreserved right after isolation or after being cultured 24 h in vitro in different preservatives.The vitality of the frozen-thawed PGCs was determined by Trypan blue exclusion method.The result showed:the vitality was the highest (87.07±1.29)% when the PGCs cryopreserved right after isolation.The thawed PGCs could form colony when cultured in vitro.The vitality of the PGCs reached highest (44.08±1.19)% when it was cultured 24 h in vitro before cryopreservation,and the PGCs could not form colony and could only survive about 40 h when culture in vitro.