昆明小鼠ES细胞的分离培养及鉴定

[目的]摸索一种适合昆明小鼠ES细胞分离培养的方法,以提高昆明小鼠ES细胞的建系率。[方法]分别采用5种方法分离ICM和ES细胞集落,摸索了以BMSCs作为饲养层时,丝裂酶素处理的合适时间。[结果]连续消化法要明显好于其他4种方法。MMC处理BMSCs 1、1.5、2 h时效果较好,3个时间点无显著差异(P>0.05)。mMEF作为饲养层与BMSCs作为饲养层时获得5代ES细胞相比差异不显著(P>0.05)。[结论]连续消化法分离ES细胞效率较高,饲养层采用MMC处理1~2 h的BMSCs饲养层或常规mMEF饲养层对昆白小鼠ES细胞无明显影响。

Isolation Culture and Identification of ES Cells in Kunming Mouse

[Objective] The purpose was to grope a method suitable for the isolation culture of ES cells in Kunming mouse so as to enhance their line-building rate.[Method] ICM and ES cell colonies were isolated by 5 methods separately and the suitable treating time of mitomycin C(MMC) with BMSCs as feeder layer was found out.[Result] The continuous digestion was better than the other 4 methods significantly.The effects of treating BMSCs by MMC for 1,1.5 and 2.0 h were better and there was no significant difference among the three time points(P>0.05).With mMEF and BMSCs as feeder layers,among the obtained 5th generation ES cells,there was no significant difference(P>0.05).[Conclusion] The continuous digestion was higher efficient in isolating ES cells.Whether the feeder layer was BMSCs treated by MMC for 1～2 h or was the conventional mMEF,it had no significant influence on ES cells in Kunming mouse.