转pGH基因猪与非转基因猪成纤维细胞系的建立及鉴定  

The Establishment and Identification of Fibroblast Cell Lines with pGH Transgenic Pigs andnon-transgenic pigs

作　　者：崔小荣[1] 李晓玲[1] 于光辉[1] 魏成晓 赵玲玲[1] 巩建飞 张廷荣[1] 孙金海[1] 

机构地区：[1]青岛农业大学动物科技学院,山东青岛266109

出　　处：《中国兽医杂志》2017年第6期3-7,I0003,共6页Chinese Journal of Veterinary Medicine

基　　金：国家转基因生物新品种培育科技重大专项(2016ZX08006-003);山东省现代农业产业技术体系生猪创新团队建设项目(SDAIT-08-13)

摘　　要：本试验采用组织块贴壁法对初生仔猪的耳组织进行原代培养,成功分离出转pGH基因猪与非转基因猪成纤维细胞,并对细胞进行形态学观察,冻存前和复苏后细胞活力检测,生长动力学分析,波形蛋白免疫组化,染色体计数以及微生物污染检测等生物学特性分析。结果表明,原代细胞经过胰蛋白酶消化和差速离心分离培养出成纤维细胞,7组细胞冻存前细胞活力均在为92%以上,冻存3个月后细胞复苏活力仍在89%以上;细胞生长总趋势呈"S"型,即经历了潜伏期、指数生长期和平台期3个阶段;细胞波形蛋白免疫组化在成纤维细胞中呈阳性反应;染色体计数结果表明,染色体数量稳定;成纤维细胞的细菌、真菌、病毒和支原体检测均为阴性。本试验成功建立了转pGH基因猪与非转基因猪成纤维细胞系。In this study,we adopted the method of explants cell culture for the newborn piglets＇ ears to the primary culture, isolated pig fibroblasts cells from pGH transgenie pigs and non-transgenic pigs. Morphological observation, determination of viability before cryopreservation and after recovery, dynamic growth analysis, vimentin immunohistochemistry, chromosome counts and microbial contamination detection were all done to study the biological characteristics of the cell lines. The results showed that the fibroblasts were cultured and isolated successfully by trypsin digestion and differential eentrifugation. The viability of seven cell lines was above 92% and 89% before eryopreservation and after recovery, respectively. The growth curves were sigmoid, and experienced the incubation period, exponential growth period and platform period. The vimentin immunohistoehemistry was positive. The chromosome counting showed that the number of chromosomes was stable. The microbial contamination detection was negative. The results indicate that the fibroblast cell lines of pGH transgenie pigs and non - transgenic pigs are successfully established.

关 键 词：pGH基因 转基因猪 成纤维细胞系 细胞培养 生物学特性 

分 类 号：S828[农业科学—畜牧学]