鸡精原细胞分离纯化与体外培养初探

Isolation,Purification and Culture of Spermatogonia in Chicken

本研究以组合酶分步消化法并利用Percoll梯度分离、贴壁纯化的方法,从不同时期的鸡睾丸组织中分离精原细胞。结果发现:①Percoll分离后,出壳6d雏鸡获得的细胞总数、精原细胞总数、精原细胞纯度分别为282.1、174.6、61.9%,其中精原细胞纯度比孵化15d、19d鸡胚和出壳后13d雏鸡分别高10.2%、5.7%和2.5%;②贴壁纯化后精原细胞纯度达到82%,比未经贴壁纯化的精原细胞纯度高出15.6%;③在无饲养层培养条件下,比较Percoll梯度分离前、后鸡精原细胞体外培养的情况,Percoll梯度分离前精原细胞贴壁在时间上比分离后较快,且分离前精原细胞体外存活时间亦比分离后长。

The study was carried out on isolation, purification and culture of spermatogonia gained from chicken testicular tissues. After enzymatic digestion, the single cell suspension was treated by percoll density gradient centrifugation to obtain high purity＇s spermatogonia. The results showed that （1）After percoll gradient centrifugation, the purity of spermatogonia gained from 6d- old-cock was 10. 2%, 5.7% and 2.5% higher than the purity from 13d-old-cock, 15d chicken embryo and 19d chicken embryo. （2）By further using adhesion purification, 82% of spermatogonia was also obtained, which was 15.6% higher than the purity of only using percoll density gradient centrifugation. （3）Before and after percoll density gradient centrifugation, the different methods for cultivation spermatogonia in vitro were also compared. Results showed that： in the condition of no CEF feed layer existed, the adhesion time and survival time of spermatogonia before percoll density gradient centrifugation was earlier and longer than the time after percoll density gradient centrifugation.