作　　者： (王艳萍）; (高帅）; (刘迥）; (曾维斌）;

机构地区： 石河子大学动物科技学院,石河子832003 塔里木大学动物科学学院,新疆生产建设兵团塔里木畜牧科技重点实验室,阿拉尔843300

出　　处： 《中国畜牧兽医》 2017年第8期2255-2260,共6页

摘　　要： 试验旨在比较不同培养方法对驴皮成纤维细胞培养的效果,以便建立其体外高效快速的培养体系。采集6~7岁健康的新疆驴真皮组织,分别用组织块贴壁法和双酶消化法对驴皮成纤维细胞进行原代培养,并检测分析细胞的生长特点、生长速度、细胞周期与支原体污染等指标。结果显示,双酶消化法(0.25%胰酶处理1h,再用0.5%胶原蛋白酶Ⅰ处理6h)培养驴皮组织3d后,成纤维细胞进入了对数增殖期,而组织块贴壁法则需要15d;细胞周期经流式分析发现,处于G0/G1期与S+G2+M期的细胞均约为50%,其余5.17%细胞处于凋亡期,表明大多细胞处于分裂增殖的活跃期,细胞具有很强的活力;试验中培养的细胞均无支原体污染。综上,应用双酶消化法可快速、高效地建立驴皮成纤维细胞体外培养体系。 This study was aimed to compare different culture methods for donkey skin fibroblasts to establish a highly efficient and rapid culture system.The dermal tissue of the healthy and 6-7 year old Xinjiang donkey was cultured for primary cells of skin fibroblast with tissue explants adherent method and double enzyme digestion method.Then,the growth characteristics,proliferation,cell cycle and mycoplasma contamination were detected and analyzed.The results showed that the fibroblast cells entered the logarithmic growth phase after cultured for 3 days by double enzyme digestion（0.25% trypsin digestion for 1hand then 0.5%collagenaseⅠfor 6h）,while it required 15 days with tissue explants adherent method.Flow cytometry analysis showed that nearly half of the cells were in G0/G1 stage,the other half in S＋G2＋M stage,while just 5.17%of the cells were apoptosis,which indicated that most of the cells were in the active stage of division and proliferation,and had strong vitality.Moreover,there was no mycoplasma contamination in the cultured cells.In summary,the culture system of donkey skin fibroblasts was rapidly and effectively established with the double enzyme digestion method in vitro.

关 键 词： 驴皮 成纤维细胞 组织块贴壁法 双酶消化法