Abstract: A new method of deriving ES cell lines from C57BL/6J mice was described. One C57BL／6J ES cell line (MESPU 41) was established in medium conditioned by rat myocardial cells, without the use of feeder cells and Leukemia Inhibitory Factor(LIF). The frequency of establishment was 1.0%. MESPU 41 cells was XX type. 89% of them showed karyotypically normal and stable which were rare in XX type ES cells. They can differentiate into wide types of cells and form teritocarcinoma in vivo. Chimeras were made by injecting the ES cells into blastocysts, indicating that they had the potency to participate into the development of embryos. Using RT-PCR method, we found that rat myocardial cells expressed LIF mRNA, which might suppress ES cells differentiation and stabilize X chromosome. In order to simplify the process of preparing conditioned medium, we also immortalized rat myocardial cells. These methods broadened the conditions for the establishment of ES cell lines and open up a new point to study the stability of cultured ES cells.

Key words: ES cell line, C57BL/6J strain, conditioned media of rat myocardial cells, chimera, immortalization

摘要： 报道一种新的建立C57BL/6J小鼠ES细胞系的方法。采用大鼠心肌条件培养基，在不使用饲养层细胞和白血病抑制因子(LIF)的情况下，从C57BL/6J品系小鼠中建成1个ES细胞系即MESPU 41，成系率为1.0%。MESPU 41细胞为XX型，核型正常率高达89%，表现出XX型ES细胞系少有的稳定性。进行体内分化实验时MESPU 41细胞能发生广泛分化形成畸胎瘤。嵌合体制作实验证实MESPU 41细胞具有嵌合能力，能参与胚胎的发育。采用RT-PCR方法，检测出大鼠心肌细胞有LIF mRNA的表达，这可能与其条件培养基保持ES细胞未分化状态并使X染色体稳定有关。同时，还对大鼠心肌细胞进行了永生化的尝试，共得到了4个永生化克隆，这将进一步简化ES细胞建系和培养工作，为进一步研究ES细胞在体外培养过程中的稳定性开创了新的起点。

关键词: ES细胞系, C57BL/6J小鼠品系, 大鼠心肌条件培养基, 嵌合鼠, 永生化