奶山羊睾丸组织冷冻保存及复苏后精原干细胞的分离培养  

Isolation and culture of spermatogonial stem cells(SSCs) from cryopreserved dairy goat testicular tissues

作　　者：朱海鲸[1] 华进联[1] 宋文聪[1] 黄建升[1] 田泽华[1] 

机构地区：[1]西北农林科技大学动物医学院陕西省干细胞工程技术研究中心农业部动物生殖发育与胚胎工程重点开放性实验室陕西省农业分子生物学重点实验室,陕西杨凌712100

出　　处：《中国兽医学报》2011年第4期583-588,593,共7页Chinese Journal of Veterinary Science

基　　金：国家自然科学基金资助项目(30972097);教育部重点科研项目(109148);中国博士后基金资助项目(20080431253);中国博士后特别科学基金资助项目(200801438);奶山羊良种繁育及产业化技术体系建设项目(nyzx07-037);国家生命科学与技术人才培养基地大学生创新性实验计划资助项目

摘　　要：以关中奶山羊为材料,比较了2种玻璃化冷冻法和1种慢速冷冻法对睾丸组织保存的效率,复苏后发现3种冻存方法均有较高的成活率(超过50%),其中玻璃化Ⅱ组达65%以上,培养及检测后均有精原干细胞存活,在体外培养可以形成胚胎干细胞样克隆,其表达精原干细胞和多能性干细胞的相关特异性标记:碱性磷酸酶(AP)、CD49f、CD133、C-Myc、Oct4和Klf4阳性,证明睾丸组织冷冻可以提供一种有效的奶山羊精原干细胞的保存方法。试验中所使用的低温保存方法简便易行,效果良好,表明这些方法可为癌症治疗等造成的无精症及少精症等不育症患者的医治以及一些珍稀濒危物种和优良畜禽的生殖细胞保存提供一种简捷有效的途径。The aim of this study was to explore an efficient procedure to cryopreserve dairy goat testis tissue.The testicular tissues of Guanzhong dairy goat were cryopreserved by two kinds of vitrification methods and a slow freezing method and compared the survival rate after thawing.The results showed high recovery rate（more than 50%）,vitrification（Ⅱ） showed high efficiency（up to 65%）,and spermatogonial stem cells（SSCs） were sucessfully obtained in these three cryoprservation methods.The cells growed and formed embryonic stem cells-like cells in vitro cultrue,and expressed AP,CD49f,CD133,C-Myc,Oct4 and Klf4.These cells can differentiate into embryonic bodies（EBs）including at least three germ layers and germ cells.This proved that testicular tissue frozen is a good route to efficiently preserve spermatogonia.It offers an efficient way to preserve SSCs for insterility and rare animals and eminent livestock.

关 键 词：睾丸组织 低温保存 精原干细胞 奶山羊 

分 类 号：S814[农业科学—畜牧学]