肝特异性表达hNPC1L1转基因巴马小型猪的获得  

Generation of transgenic Bama minipigs with liver-specific expression of human Niemann-Pick C1-like 1(hNPC1L1)

作　　者：许崇利 许崇波[2] 宫语晨 欧阳红生 

机构地区：[1]吉林化工学院生物与食品工程学院,吉林吉林132022 [2]韶关学院英东生命科学院,粤北生猪生产及疫病防控协同创新发展中心,广东韶关512005 [3]吉林农业大学动物科学技术学院,吉林长春130118 [4]吉林大学动物医学学院,吉林长春130062

出　　处：《江苏农业学报》2017年第5期1069-1075,共7页Jiangsu Journal of Agricultural Sciences

基　　金：国家科技支撑计划项目(2011BA115B02)

摘　　要：利用Fu GENE6将线性化的p Liv-11-Neor/Kanr-h NPC1L1转染至小型猪胎儿成纤维细胞,G418筛选和PCR鉴定结果表明,成功获得了肝特异性表达人尼曼-匹克C1型类似蛋白1(Human Niemann-Pick C1 Like 1,h NPC1L1)转基因小型猪胎儿成纤维细胞。利用体细胞核移植技术将转h NPC1L1基因小型猪胎儿成纤维细胞作为核供体细胞,获得了肝特异性表达h NPC1L1转基因巴马小型猪,并对转基因巴马小型猪进行了基因组水平、转录水平和表达水平检测,同时对目的基因进行了免疫组化定位分析。结果显示,目的基因特异性地表达于肝组织,并且定位于肝小叶间胆管膜上,表明已成功获得了转h NPC1L1基因巴马小型猪,从而为研究尼曼-匹克C1型类似蛋白1(Niemann-Pick C1 Like 1,NPC1L1)在肝中作用的分子机制提供了很好的模型动物。In this study,the minipig fetal fibroblast cells with human Niemann-Pick C1-like NPC1L1 specially ex-pressed in liver were obtained through transfecting linearized vector pL／v-11-Neor/K a n r-hN P C lL l into minipig fetal fibro-blast cell by FuGENE ？ 6. The positive clones were selected with G418 and identified by PCR. The transgenic Bama mini-ature pigs with hNPC1L1 specifically expressed in the livers were generated by using somatic cell nuclear transfer technique with the minipig fetal fibroblast cells as nuclear donor cells. The transgenic Bama miniature pigs were detected on the ge-nomic level,transcription level and expression level. Meanwhile,the target gene was analyzed by immunohistochemistry.The results showed that the target gene was specifically expressed in the liver tissue and localized on the hepatic interlobular hNPC1L1 transgenic Bama miniature pigs will provide a model animal for studying the molecular mechanism ofNPC1L1 in liver.

关 键 词：人尼曼-匹克C1型类似蛋白1(hNPC1L1) 肝特异性表达 体细胞核移植 巴马小型猪 

分 类 号：S858.286.5[农业科学—临床兽医学]