3个不同品系小鼠ES细胞的绿色荧光蛋白标记(英文)  

Labeling of Three Different Mouse ES Cell Lines with the Green Fluorescent Protein

作　　者：赵文宁[1] 孟国良[1] 薛友纺[1] 

机构地区：[1]北京大学生命科学学院,北京100871

出　　处：《Acta Genetica Sinica》2003年第8期743-749,共7页

基　　金：国家自然科学基金资助项目 (编号 :3 0 0 70 414 )~~

摘　　要：用电穿孔法将线性化的质粒pEGFP N3分别导入来自 12 9/ter、C5 7BL/ 6J和BALB/c 3个品系的小鼠胚胎干细胞MESPU 13、MESPU 35和MESPU 6 2中 ,经G4 18筛选、荧光显微镜镜检、阳性克隆扩增、流式细胞仪分选、再扩增以及核型分析等过程 ,分别得到核型大于 85 %的被EGFP稳定标记的细胞株 5个 (12 9/ter 2个、C5 7BL/ 6J 1个、BALB/c 2个 ) ,分别命名为MESPU 13/G1和MESPU 13/G2、MESPU 35 /G1、MESPU 6 2 /G1和MESPU 6 2 /G2。从不同品系中各选一个增殖生长快、形态典型的标记细胞株 ,进行碱性磷酸酶染色、oct4基因产物的表达检测、类胚形成和体内分化鉴定 ,结果表明所得到的核型正常的、稳定标记的ES细胞系具有原ES细胞的典型特征。The linearized plasmid pEGFP-N3 was electroporated into three different mouse ES cell lines MESPU-13,MESPU-35 and MESPU-62 derived from 129/ter,C57BL/6J and BALB/c mouse strains respectively.Resistant clones were selected in the presence of G418 and then were identified under the fluorescence microscope through blue exciting light.Positive green clones were primarily expanded and further sorted using FACS(fluorescence activated cell sorter).Finally five EGFP stable integrated cell strains were obtained and were expanded (2 strains from 129/ter,1 strain from C57BL/6J and 2 strains from BALB/c).Each of the five cell strains presents high proliferation growth rate and typical morphology characters of ES cells and their colonies.More than 85% cells of each cell strain contain normal diploid karyotype.Then some analysis such as the AP (alkaline phosphatase) staining, oct4 gene expression assay,embryonic body formation and differentiated test in vivo and in vitro were made.The results indicated that the stable labeled ES cell strains had the normal karyotypes and maintained the ES cell typical characteristics.

关 键 词：小鼠胚胎干细胞 绿色荧光蛋白 电穿孔 体内分化 

分 类 号：Q952[生物学—动物学]