VEGF165真核表达载体的构建及其在鼠膀胱平滑肌细胞的表达

目的 构建血管内皮生长因子(VEGF165)真核表达载体pcDNA3.1(-)/VEGF165,转染鼠膀胱平滑肌细胞并检验其生物学活性.方法 将VEGF165 cDNA克隆于真核表达载体pcDNA3.1(-),构建真核表达载体pcDNA3.1(-)/VEGF165;并转染入鼠膀胱平滑肌细胞,采用RT-PCR和免疫荧光染色检测VEGF165基因在鼠膀胱平滑肌细胞中的表达,并用MTT法检测转染后细胞上清液中VEGF165的生物学活性.结果 和结论将构建的真核表达载体pcDNA3.1(-)/VEGF165转染入鼠膀胱平滑肌细胞后,VEGF的表达增高,转染后细胞上清液具有促使内皮细胞增殖的生物学活性.

Construction of eukaryotic expression vector for VEGF165 gene and its expression in rat bladder smooth muscle cells

Objectives To construct an eukaryotic expression vector for vascular endothelial growth factor(VEGF)165 gene and obtain VEGF expression in rat bladder smooth muscle cells.Methods VEGF165 cDNA was cloned into the eukaryotic expression vector pcDNA3.1(-),and the resultant recombinant vector pcDNA3.1(-)/VEGF165 was transfected into the rat bladder smooth muscle cells by electroporation.VEGF expression in the cells was determined by RT-PCR and immunofluoresence assay,and the biological activity of VEGF in the supernant of the transinfected cell culture was tested by MTT assay.Results and conclusion VEGF expression was obtained in the transinfected cells,and the supernant of the transinfected cell cultures stimulated the proliferation of the endothelial cells.