体细胞核移植生产梅山猪  

Production of Cloned Meishan Piglets by Somatic Cell Nuclear Transfer

作　　者：华再东[1] 郑新民[1] 魏庆信[1] 肖红卫[1] 刘西梅[1] 华文君[1] 郭帅 张立苹[1] 毕延震[1] 任红艳[1] 岑文[3] 华升 

机构地区：[1]湖北省农业科学院、动物胚胎工程及分子育种湖北省重点实验室,湖北武汉430064 [2]武汉金鹰牧业有限公司,湖北武汉430209 [3]陕西师范大学生命科学学院,陕西西安710119

出　　处：《中国畜牧兽医》2014年第10期192-197,共6页China Animal Husbandry & Veterinary Medicine

基　　金：国家转基因生物新品种培育重大专项(2014ZX08010-003、2014ZX08006-003);湖北省农业科技创新中心课题(2011-620-001-003);湖北省自然科学基金重点课题(2010CDA120)

摘　　要：本研究以中国优良地方品种梅山猪为材料,采用胰酶消化法获得猪胎儿成纤维细胞,通过使用Y染色体SRY基因引物SRY-1、SRY-2进行PCR扩增,结果发现,雄性胎儿样本能扩增出250bp的Y染色体特异性基因片段,而雌性胎儿样本未扩增出此条带;G1、G2、G3、G4和G5代的融合效率差异不显著(P>0.05),但G5代作为供体细胞核移植的重组胚胎卵裂率显著高于G1、G2、G3和G4代(P<0.05);使用G5代的胎儿成纤维细胞作为供核细胞,通过手术移植的方法,将重构胚胎移植到二元后备母猪体内,结果成功地获得了体细胞克隆梅山仔猪,并且仔猪全部为雄性胎儿,说明该性别鉴定方法不但操作简单,而且准确度高;经微卫星DNA多态性鉴定,确定克隆猪来自供核细胞,与代孕母猪无亲缘关系。本研究将为中国优质猪品种改良、保种、性别控制及建立人类疾病模型等研究提供有效可行的方法,为批量生产克隆优秀种猪提供了坚实的理论基础。In this study,Meishan pigs of Chinese excellent local varieties was used as materials,the pig fetal fibroblasts was obtained by pancreatic enzyme digestion method,SRY-1and SRY-2were used as primers,though PCR identification,male fetus samples could amplify out 250 bp of Y chromosome specific genes,while the female fetus samples could not amplify the stripe.The fusion efficiency of nuclear transfer embryo using G1,G2,G3,G4 and G5generation as a donor cell had no significant difference（P〉0.05）,but the cleavage rate by SCNT using G5 generation cell was significantly higher than G1,G2 and G3and G4（P〈0.05）.Used the G5 generation of fetal fibroblasts of Meishan pigs as donor cells,we successfully cloned a Chinese Meishan pig by SCNT,and the piglets were all male fetus.The results showed that the method of sex identification was not only the easy operation,but also high accuracy,the analysis of microsatellite genotyping for Meishan pigs confirmed that the piglets were genetically derived from fetal fibroblasts of Meishan pigs and unrelated to their surrogate mother.This study will provide a effective feasible method for pig breeding,gender control,and human disease models,and lay a foundation for mass production of cloned excellent breeding with high productivity.

关 键 词：梅山猪 克隆猪 核移植 猪胎儿成纤维细胞 

分 类 号：Q813[生物学—生物工程]