潮霉素抗性的小鼠胚胎干细胞滋养层的制备

目的：获得潮霉素抗性的小鼠胚胎干细胞滋养层。方法：利用基因组整合了潮霉素B磷酸转移酶基因的小鼠品系Smad4hygro＋/-,从受精14d的小鼠胚胎中制备原代胚胎成纤维细胞;鉴定基因型后,挑选一株潮霉素B磷酸转移酶基因杂合型胚胎成纤维细胞,用丝裂霉素C处理,以获得潮霉素抗性的滋养层细胞。结果：经潮霉素B加压处理后,杂合型滋养层细胞可存活2周左右,并可维持胚胎干细胞的生长。结论：制备的滋养层细胞可用于潮霉素抗性胚胎干细胞的筛选。

Preparation of Hygromycin Resistant Fibroblast Feeder Layer Cells for Mouse Embryonic Stem Cells

Objective： To prepare a stock primary mouse embryomic fibroblast （MEF） cells with hygromycin resistance. Methods： Female mice containing heterozygous hygromyein B phosphotransferase gene were sacrificed 14 days after coitus to get the embryos, MEF cells were isolated and genotyped by PCR. Cells containing hygromycin B phosphotransferase gene were chosen and treated with mitomycin C to obtain the feeder layer cells. Results： After treatment with hygromycin B medium, the feeder layer cells could survive up to about 2 weeks and support the growth of the embryonic stem （ES） cells. Conclusion： The results demonstrated that the hygromycin resistant fibroblast feeder layer cells could be used to screen the hygromycin resistant ES cells.