文档介绍：海人藻酸受体亚基GluR6特异性shRNA真核表达载体的构建
【摘要】目的构建海人藻酸受体亚基GluR6的shRNA真核细胞表达载体,进一步研究GluR6在脑缺血中的作用机制。方法根据GenBank数据库中GluR6的cDNA序列设计靶序列,化学合成两条互补的DNA寡核苷酸链,连接质粒后转染大肠杆菌,使其在大肠杆菌内大量复制;提取质粒,进行限制性内切酶酶切鉴定及测序分析。结果限制性内切酶酶切鉴定及测序分析证实了序列的正确性,与合成的寡核苷酸序列完全相符,且以正确的方向插入。结论成功构建了GluR6特异性shRNA真核表达载体。
【关键词】 GluR6 载体
Abstract:Objective To construct the eukaryotic expression plasmid for kainate receptor (KA) subunit GluR6 specific According to the GluR6 cDNA sequence, two strands of oligonucleotide were designed and chemically synthesized, and then annealed into two double strands in vitro. The DNA strands were ligated with plasmid vector pSilencer-GluR6. The binate plasmid was transfected into E. coli and multiplied. After being extracted from E. coli, the plasmids were processed to verify the sequence of the shRNA by restriction endonuclease digestion and The binant plasmid was found containing correct plete sequence of the shRNA, indicating that the specific GluR6 shRNA had
been inserted into the eukaryotic expression The eukaryotic expression plasmid pSilencer-GluR6 has been essfully constructed.
Key words: GluR6; shRNA; vector
谷氨酸(Glu)S) 中重要的神经递质,S,S 的突触传递过程中发挥着重要作用。另外,Glu能神经传递的异常往往与一些神经疾病发生有关,如癫疒间、缺血、阿尔茨海默病等[1-2]。Glu受体分为代谢型Glu受体(mGluR)和离子型Glu受体(iGluR)。 iGluR是非特异性阳离子通道,根据它们所首选的配体的不同可分为 NMDA 受体和非NMDA受体两类,非NMDA受体又可分为AMPA受体和KA受体[3-4]。由于缺乏选择性药物,KA受体在