期刊论文

Construction and Expression of Eukaryotic Vector of FXR1 and the Effection of Ganglioside GM1

中文

现代生物医学进展

1673-6273

2014

14

28

5407-5409,5435

10.13241/j.cnki.pmb.2014.28.002

国家自然科学基金青年项目（81200881） 湖南省自然科学基金项目（12JJ6073）

本校为第一机构

目的:构建脆性X相关基因1(FXR1)的真核表达载体并检测其对神经节苷脂(GM1)浓度的影响.方法:以pYESTrp3-FXR1为模板,利用PCR扩增FXR1基因,PCR产物经EcoRⅠ和XhoⅠ双酶切后插入真核表达载体pcDNA3.1(-)中,获得的阳性克隆进一步酶切及测序鉴定;将构建成功的pcDNA3.1 (-)-FXR1转染SH-SY5Y细胞后,采用Western blot检测FXR1的表达情况,同时采用ELISA试剂盒检测细胞内GM1的浓度.结果:PCR扩增产物为1.9 Kb的片段,与FXR1基因大小相符,阳性克隆经双酶切后获得两条分别为5.4 Kb和1.9 Kb的片段,测序结果与GeneBank中序列相同.构建成功的重组质粒pcDNA3.1 (-)-FXR1转染SH-SY5Y细胞后,细胞中FXR1的表达增加,同...

Objective： To construct a eukaryotic expression vector encoding FXR1 and detect its effect on the concentration of GM1. Methods： FXR1 was amplified by PCR using pYESTrp3-FXR1 as template and then inserted into eukaryotic expression vector pcDNA3.1 （-） using restriction site ofEcoR I and Xho I. The positive clones were further identified by enzyme cleaving and sequencing. The pcDNA3.1 （-）-FXR1 was transfected into SH-SY5Y cells and the expression level of FXR1 was detected by Western blotting and the concentration of GM1 was detected by EL...