Cloning and analysis of the genes during the induction of mouse embryongenesis

将小鼠胚胎干细胞（ES细胞）与巨噬细胞集落刺激因子缺乏的OP9基质细胞共培养，可通过中胚层细胞分化为造血细胞。为了克隆在这一过程中起作用的细胞外信号蛋白基因，利用第5天分化诱导细胞进行cDNA克隆。利用信号序列诱捕法（Signal Sequence Trap， SST）克隆秘书蛋白和I型跨膜蛋白是本课课组创新的一种高效克隆方法。因此，一个cDNA片段必须包含真实的信号序列。以SST cDNA片段为探针克隆全长cDNA，该候选基因是鸡基因的鼠同源基因，其表达受c-Myb控制，但功能尚不清楚。目前，对该基因的表达和功能进行了研究。同时，我们认为通过研究胚胎干细胞与原始生殖细胞基因表达的异同，可以克隆诱导小鼠早期胚胎发生的相关基因。我们开始从纯化的原始生殖细胞中收集mRNA。我们计划进行差异显示或SAGE（基因表达序列分析）克隆基因。

When mouse embryonic stem cells (ES cells) are co-cultured on macrophage colony stimulating factor deficient OP9 stromal cells, hematopoietic cells can be differentiated via mesodermal cells. In order to clone the genes of extraccellular signaling proteins functioning during this process, cDNA cloning was carried out by using the day 5 differentiation induced cells. We utilized Signal Sequence Trap (SST) method which is an efficient cloning strategy for secretary proteins and type I transmembrane proteins and the method was innovated by our group.As a result, one cDNA fragment which must contain authentic signal sequence. Full length cDNA was cloned by using the SST cDNA fragment as aprobe, this candidate gene turns out to be a murine homologue of the chicken gene whose expression is controlled by c-Myb but whose function is unknown yet. Now, the expression and the function of this gene is investigated.Meanwhile, we consider it possible to clone the genes involved in the induction of early embryogenesis of mice by examining the similarities and the differences of the gene expression between ES cells and primordial germ cells. And we start to collect mRNA from purified primordial germ cells. We are planning to carry out differential display or SAGE (serial analysis of gene expression) to clone the genes.