长牡蛎精子超低温冷冻后超微结构损伤研究  

STUDY ON ULTRASTRUCTURE OBSERVATION OF FRESH AND CRYOPRESERVATED SPERM IN CRASSOSTREA GIGAS

作　　者：韩龙江 刘清华[1,3,4] 许飞[1,3,4] 温海深[2] 李军[1,3,4] 

机构地区：[1]中国科学院海洋研究所,中科院实验海洋生物学重点实验室,青岛266071 [2]中国海洋大学,青岛266009 [3]海洋生态养殖技术国家地方联合工程实验室,青岛266071 [4]青岛海洋科学与技术国家实验室,海洋生物学与生物技术功能实验室,青岛266237

出　　处：《水生生物学报》2017年第1期220-227,共8页Acta Hydrobiologica Sinica

基　　金：国家高技术研究发展计划(863计划)项目(2012AA10A402);海洋经济创新发展区域示范项目(12PYY001SF08);国家重点基础研究发展计划(973计划)项目(2010CB126401);鳌山科技创新计划(2015ASKJ02,2015ASKJ02-03-03);水产种质资源平台运行服务项目;中国科学院青促会项目资助~~

摘　　要：采用程序降温仪分步降温冷冻保存长牡蛎(Crassostrea gigas)精液,并用扫描电镜、透射电镜研究了精子的超微结构损伤。超低温冷冻保存后长牡蛎精子的运动率、受精率及孵化率与鲜精无显著差异。鲜精中84.5%的精子形态结构正常,冻精中73%的精子形态结构正常。形态结构正常的精子表现为顶体、质膜、线粒体与鞭毛结构完整、染色质形状规则,顶体、线粒体及中心粒结构正常,鞭毛形态完整、微管结构清晰;形态结构异常的精子表现为顶体脱落、解体,精子头部质膜膨胀、破裂、染色质肿胀、破裂、解体,线粒体移位、脱落、膨胀,嵴退化或消失,鞭毛弯折、断裂,微管解聚。结果显示,以10%DMSO为抗冻保护剂,HBSS溶液为稀释液,1鲶4的稀释比例,添加海藻糖,采用分步降温法冷冻保存,对长牡蛎精子具有较好的抗冻保护作用,合适的冻存方法可以有效的保护太平洋牡蛎精子冷冻过程中结构损伤。研究有助于长牡蛎种质资源的收集保存及应用。A step-wise cooling schemes was employed to cryopreserve Crassostrea gigas sperm, and the sperm ultra-structure was observed by scanning electron microscopy and transmission electron microscopy. The results showed that there were no significant differences between frozen-thawed sperm and fresh sperm in the motility, fertilization rate and hatching rate. Both the fresh sperm and cryopreserved sperm had ultrastructural damages. The normal rates of the fresh and cryopreservated sperms were 84.5%and 73%, respectively. The cryopreserved sperm without damage had normal morphology in the plasma membrane, mitochondria and nuclear, the acrosome, and centriole, and the mitochondrion obtained integrity with well-developed cristae. The sperm cryodamages with damages had swelled or disrupted plasma and nuclear membrane, partially damaged nucleus and swelled, dislocated or disarticulated mitochondrion with dege-nerated or vanished cristae. The results showed that HBSS with at 1 4 dilution trehalose and 10%DMSO is the best condition for extender and cryoprotecant and for protecting the frozen-thawed C. gigas sperm, which will benefit the preservation of C. gigas and application of sperm cryopreservation skills.

关 键 词：长牡蛎 精子 扫描电镜 透射电镜 超微结构 

分 类 号：S917[农业科学—水产科学]