胰岛素样生长因子-1逆转录病毒表达载体的构建及其在大鼠胚胎神经干细胞中的表达  

Construction of retroviral vector containing insulin like growth factor-1 and its expression in rat neural stem cells

作　　者：王进[1] 王红星[1] 崔景彬[2] 

机构地区：[1]周口师范学院生命科学系,周口466000 [2]北京市神经外科研究所,北京100050

出　　处：《郑州大学学报（医学版）》2007年第6期1136-1140,共5页Journal of Zhengzhou University(Medical Sciences)

摘　　要：目的:构建胰岛素样生长因子-1(IGF-1)逆转录病毒表达载体,并观察转IGF-1基因的鼠胚胎神经干细胞(NSC)体内外基因表达情况。方法:通过酶切和连接的方法将IGF-1基因克隆入逆转录病毒表达载体pLXSN中,感染包装细胞后收集病毒颗粒再感染从胎龄14d的SD大鼠全脑组织中分离出的NSC,RT-PCR检测IGF-1的mRNA,免疫组织化学法检测IGF-1蛋白的表达,并用立体定向法将Brdu标记的转基因NSC移植到大鼠大脑中动脉梗死模型的纹状体缺血半暗区,免疫染色观察移植后1周转基因NSC在脑内的基因表达情况。结果:酶切及序列分析证实重组逆转录病毒载体pLX SN-IGF-1构建正确,RT-PCR能检测到转基因IGF-1的mRNA,免疫组织化学法能检测到IGF-1在转染了逆转录病毒的NSC及其子代细胞内表达,转基因NSC移植后1周在宿主脑内迁徙并表达IGF-1基因产物。结论:成功构建了携带IGF-1基因的逆转录病毒载体,转染了重组逆转录病毒载体的NSC可在体内外表达IGF-1。Aim ： To construct retroviral vector containing insulin like growth factor-1 （ IGF-1 ） and observe its expression in rat neural stem cells in vivo and in vitro. Methods：The recombinant retroviral vector pLXSN-IGF-1 containing IGF- 1 gene was established using the method of enzyme cutting and ligation, The recombinant vector was transferred to packaging cell line pA317 to acquire virus, then the recombinant virus was used to infect neural stem cell（NSC） isolated from the whole brain of embryonic day 14 SD rats. IGF-1 mRNA,expression of IGF-1 were detected using RT-PCR, immunocytochemistry, respectively, Stereotactic operation was made to transplant NSC marked with Brdu to striatum ischemia penumbra of the middle cerebral artery infarction model rat. IGF-1 gene expression of transplanted cells 1 week after transplantation was detected using immuno-staining. Results：The construction of retroviral vector pLXSN-IGF-1 was successfully established. RT-PCR and immunochemistry verified the expression of IGF-1 in NSC 1 week after transplantation. Immunochemistry showed that transgenic NSC migrated and expressed IGF-1 in host brain. Conclusion： Retroviral vector pLXSN-IGF-1 had been successfully established. IGF-1 gene could express in rat NSC in vivo and in vitro.

关 键 词：神经干细胞 胰岛素样生长因子-1 逆转录病毒 转染 转基因 大鼠 

分 类 号：Q785[生物学—分子生物学]