论文部分内容阅读
EDAG是在胚胎发育阶段造血干细胞特异性表达的基因.为了在早期造血组织细胞中实现相关基因的条件敲除,构建了含有早期造血组织特异性表达的EDAG启动子和Cre重组酶基因的转基因EDAG-Cre表达载体质粒.通过显微注射的方法将线性化的5.6kb的EDAG-Cre转基因片段导入小鼠受精卵细胞核,获得的新生小鼠经过PCR鉴定,常规方法培育传代.结果发现,共获得了6只阳性转基因首建鼠,其中4只已经建系并稳定传代.RT-PCR分析表明Cre重组酶基因在阳性转基因小鼠的骨髓、脾脏、胸腺、外周血以及胎肝等组织中均有表达,重组酶活性也在脾和骨髓中获得确认.EDAG-Cre重组酶转基因小鼠的建立,为研究早期造血组织以及造血干细胞特异性基因条件敲除小鼠模型的建立奠定了基础.
EDAG is a gene specifically expressed in hematopoietic stem cells during embryonic development.For conditional knockdown of related genes in early hematopoietic tissue cells, a transgenic EDAG containing the EDAG promoter and Cre recombinase genes specifically expressed by early hematopoietic tissue was constructed -Cre expression vector plasmid.The linearized 5.6kb EDAG-Cre transgene fragment was introduced into the nucleus of mouse fertilized egg by microinjection, and the newborn mice obtained were identified by PCR and passaged by conventional methods.The results showed that a total of Six positive transgenic mice were established, of which four had been established and stably passaged.RT-PCR analysis showed that the Cre recombinase gene was expressed in the bone marrow, spleen, thymus, peripheral blood and fetal liver of the positive transgenic mice Expression and recombinase activity were also confirmed in the spleen and bone marrow.The establishment of transgenic mice with EDAG-Cre recombinase laid the foundation for the establishment of a mouse model of early hematopoietic tissue and conditional hematopoietic stem cell-specific knockout.