Neurturin基因的克隆及其在Vero细胞中的表达

Cloning of Neurturin Gene and Its Expression in Vero Cells

目的克隆Neurturin(NTN)基因,并在Vero细胞中表达。方法用RT-PCR方法扩增hNTN cDNA,并克隆至pcDNA3真核表达载体中,经Lipofectamine2000转染Vero细胞,挑选稳定表达克隆,用RT-PCR及免疫荧光分析鉴定,并对转染后细胞的形态及生长状况进行分析。结果重组质粒pcDNA3/hNTN转染Vero细胞后获得了稳定表达克隆,且有目的蛋白表达,转染后细胞形态和生长特性发生了一定改变。结论获得了稳定表达NTN蛋白的Vero细胞株,为其移植并进行帕金森病猴基因治疗动物实验奠定了基础。

Objective To clone Neurturin （NTN） gene and express in Vero cells.Methods Amplify human NTN eDNA by RT-PCR and clone to eurkaryotie expression vector peDNA3. Transfect Vero cells with the constructed recombinant plasmid peDNA3/hNTN in mediation of Lipofectamine 2000. Identify the expressed product by RT-PCR and IFA. Observe the morphology and growth of transfected Vero cells. Re- sults The positive clones for stable expression of NTN was obtained after transfection of Vero cells with recombinant plasmid peDNA3/hNTN, and the target protein was expressed. The morphology and growth of transfected Vero cells showed significant change. Conclusion The Vero cell strain for stable expression of NTN was established, which laid a foundation of gene therapy of Parkinson disease in rhesus.