多巴胺D1受体RNA干扰载体的构建及其沉默效应鉴定  

Construction of RNAi vector of dopamine D1 receptor and identification of its silencing effects

作　　者：李辉[1,2] 徐军美 袁贵秀[1,2] 李晋[1,2] 陈章玲[1,2] 

机构地区：[1]中南大学湘雅二医院麻醉科 [2]中南大学麻醉医学研究所,长沙410011

出　　处：《中南大学学报（医学版）》2013年第6期570-575,共6页Journal of Central South University ：Medical Science

摘　　要：目的:构建多巴胺D1受体(DRD1)表达干扰载体,为研究DRD1在神经细胞中的作用及抗惊厥药物打下基础。方法:根据GenBank中DRD1基因序列,设计10个干扰序列并构建DRD1干扰载体。对NG-108-15进行脂质体转染后,GFP标记检测转染效果。Real-time PCR和Western印迹检测NG-108-15中DRD1的表达量。结果:构建的10个干扰载体均能采用脂质体法转染到NG-108-15细胞中。转染后NG-108-15中DRD1 mRNA和蛋白的表达均明显下降。其中转染pGPU6-GFP-Neo-si-DRD1-5载体的NG-108-15中DRD1 mRNA表达水平最低,而转染pGPU6-GFP-Neo-si-DRD1-1,-2,-6,-7载体的NG-108-15中DRD1蛋白表达水平最低。结论:成功构建了DRD1表达干扰载体。Objective： To construct dopamine D1 receptor （DRD1） expression interference vectors to study the role of DRD1 in nerve cells and lay a foundation for drug development in anti-convulsion. Methods： Based on DRD1 gene sequence in GenBank, 10 interfere vectors of DRD 1 were designed. Liposomal was used to transfect NG-108-15 and the transfect effect was assayed by GFP. With realtime PCR and Western blot, the DRD 1 expression was detected. Results： The 10 constructed interfere vectors transfected into NG-108-15 cells by liposomal method and inhibited DRD1 mRNA and protein expression. DRD1 mRNA expression in NG-108-15 cells transfected with pGPU6-GFP-Neo-si-DRD1-5 was the lowest whereas DRD1 protein expression in NG - 108-15 cells transfected with pGPU6-GFP-Neo-si-DRD 1 - 1, -2, -6, -7 was the lowest. Conclusion- DRD 1 expression interference vector is successfully constructed.

关 键 词：多巴胺D1受体 RNA干扰 NG-108-15细胞 

分 类 号：R614[医药卫生—麻醉学]