中国比较医学杂志2016，Vol.26Issue(7)：31-34,63,5.DOI:10.3969.j.issn.1671-7856.2016.07.005
真核表达载体GV394-Nurr1的构建及鉴定
The construction and identification of eukaryotic expression vector GV394-Nurr1
摘要
Abstract
Objective To construct the eukaryotic expression vector GV394⁃Nurr1 containing human Nurr1 gene and to study the effects of transient transfection of Nurr1 on intracellular reactive oxygen species level. Methods The full⁃length of human Nurr1 gene amplified by PCR was subcloned into T vector and sequenced. GV394⁃Nurr1 vector was constructed by BamHI and XhoI double digestion and then T4 DNA ligase conjunction. GV394⁃Nurr1 was transfected into SH⁃SY5Y cells by liposome transfection technique;The mRNA of Nurr1 was detected by RT⁃PCR;The effect of Nurr1 expression on intracellular reactive oxygen species ( ROS ) was detected by ( DCFH⁃DA ) staining. Results PCR and sequencing confirmed that the Nurr1 gene was correctly cloned into eukaryotic expression vector GV394. The RT⁃PCR results showed that the Nurr1 mRNA expression in the neuroblastoma SH⁃SY5Y cells transiently transfected Nurr1 was higher than that in the control group. DCFH⁃DA staining showed that the level of reactive oxygen peak in neuroblastoma cells transiently transfected Nurr1 obviously shifted to the left compared to the control group. Conclusions The human Nurr1 gene eukaryotic expression vector was successfully established and its high expression in the neuroblastoma SH⁃SY5Y cell line significantly decreased the ROS level. This provide the basis for further study on the function of Nurr1 in vitro and its relationship with the protective effect of dopaminergic neurons.关键词
Nurr1/真核表达载体/帕金森氏病/细胞活性氧Key words
Nurr1/Eukaryotic expression vector/Parkinson disease/Reactive oxygen species分类
医药卫生引用本文复制引用
樊秀双,郭军堂,陈安琪..真核表达载体GV394-Nurr1的构建及鉴定[J].中国比较医学杂志,2016,26(7):31-34,63,5.基金项目
山东省自然科学基金（ZR2011HM014）。 （）
