摘要
对绵羊颗粒细胞进行体外培养、冷冻复苏。采集西藏绵羊卵巢,用注射器抽吸卵泡中的卵泡液,经低速离心分离沉淀得到卵泡内的颗粒细胞,用于体外培养。结果显示:原代培养时,颗粒细胞形成单层需要6~7d,形状多为梭形或不规则三角形,呈放射状;传代颗粒细胞生长速度较快,接种后第二天细胞就开始贴壁,第6d左右长满平皿底,形成颗粒细胞单层,多呈流线型。当传至第3代培养后,即可获得纯化的颗粒细胞;冻存的细胞经复苏后仍保持其正常的结构和功能。细胞生长曲线呈"S"形,符合贴壁细胞的生长特征。可见,所建立的西藏绵羊颗粒细胞分离方法和保存方法可行有效,这能为西藏绵羊的体细胞克隆提供供体细胞,也能为建立西藏绵羊遗传资源体细胞库等研究奠定基础。
Sheep granulosa cells were cultured in vitro and resuscitated by freezing.The ovaries of Tibetan sheep were collected, and the follicular fluid in the follicle was aspirated with a syringe, and the granulosa cells in the follicle were obtained by low-speed centrifugation for in vitro culture.The results showed that it took 6~7 days for granulosa cells to form monolayer in primary culture.The granulosa cells in the passage grew faster, and the cells began to stick to the wall on the second day after inoculation.On the around 6d, the bottom of the dish was covered, forming a single layer of granulosa cells, which were mostly streamlined.The purified granulosa cells were obtained after culture in the third generation.After resuscitation, the frozen cells maintained their normal structure and function.The curve of cell growth was "S" shape, which accorded with the growth characteristics of adherent cells.In brief, it was feasible and effective that the methods established for isolation and preservation of Tibetan sheep granulosa cells, which would provide necessary donor cells for somatic cell cloning of Tibetan sheep and lay a foundation for the establishment of somatic cell bank of Tibetan sheep genetic resources.
作者
田孟芳
卢晓丽
程志鹏
任子利
孙磊
赵彦玲
Tian Mengfang;Lu Xiaoli;Cheng Zhipeng;Ren Zili;Sun Lei;Zhao Yanling(Animal Science College,Tibet Agriculture&Animal Husbandry University,Nyingchi Tibet,860000,China)
出处
《高原农业》
2019年第5期510-514,共5页
Journal of Plateau Agriculture
基金
西藏自治区班戈县畜牧业综合发展关键技术研究（603318014）
关键词
西藏绵羊
颗粒细胞
体外培养
生长曲线
Tibetan sheep
granular cell
in vitro culture
growth curve
