| 摘要: |
| 【目的】研究云南半细毛羊毛囊干细胞(Hair follicle stem cells，HFSCs)的分离培养方法。【方法】用组织块法原代培养云南半细毛羊HFSCs，并进行细胞纯化和传代培养，培养液为无血清培养液DMEM/F12+40 ng/mL bFGF+20n g/mL EGF+20 μL/mL B27+100 IU/mL青霉素+100 IU/mL链霉素。对建立的细胞系进行免疫组化、RT-PCR和流式细胞术鉴定。【结果】HFSCs体积小，为贴壁生长细胞，呈典型的铺路石状，核质比高，在倒置显微镜下胞体透亮、折光性强。RT PCR分析表明，HFSCs表达K14、K19和β1integrin，但是不表达CD271和nestin。免疫组化分析结果表明，HFSCs表达K14，不表达CD271。流式细胞术鉴定表明，HFSCs表达K14，阳性率为93.5%；不表达CD271。【结论】 建立了云南半细毛羊HSFCs无血清体外培养技术体系。 |
| 关键词: 云南半细毛羊 毛囊干细胞 细胞鉴定 |
| DOI： |
| 分类号: |
| 基金项目:国家现代农业产业技术体系建设项目(CARS-40)；云南省应用基础研究青年项目(2012FD083) |
|
| Establishment of hair follicle stem cell line for Yunnan semi-fine wool sheep |
|
Lü Chunrong,QUAN Guobo,WU Guoquan,et al
|
| Abstract: |
| 【Objective】This study established an isolated culture method for hair follicle stem cells (HFSCs) of Yunnan semi-fine wool sheep.【Method】The tissue piece culture method was used to separate and culture HFSCs followed by cell purification and subculture.Culture medium was serum free medium DMEM/F12+bFGF 40 ng/mL+EGF 20 ng/mL+B27 20 μL/mL+100 IU/mL penicillin+100 IU/mL streptomycin.Then,the established cell lines were identified by immunohistochemistry,RT PCR,and flow cytometry.【Result】The established HFSCs had small volume,and line exhibited adherent growth type,high nucleus/cytoplasm ratio,high refractivity,and transparent cell appearance.RT-PCR indicated that HFSCs expressed K14,K19,and β1integrin but not CD271 and nestin.The immunofluorescence data indicated that HFSCs expressed K14 but not CD271.The flow cytometry indicated that HFSCs expressed K14 with positive rate of 93.5%,but not CD271.【Conclusion】This study established the in vitro non serum culture procedure and cell identification technology for HFSCs of Yunan semi fine wool sheep. |
| Key words: |
