Neurturin基因的克隆及其在C17.2神经干细胞中表达的研究

Study of neurturin gene clone and its expression in C17.2 neural stem cell

目的 探讨Neurturin(NTN)基因转染的C17.2神经干细胞移植后对帕金森病(PD)大鼠模型的保护作用,为其提供基因修饰的神经干细胞。方法 用 RT-PCR方法获取Prepro-NTN cDNA,并克隆至 pBlueskiptⅡ载体中,再将 Prepro-NTN基因克隆至 pcDNA3.1-hygro真核表达载体中,经 Lipofectamine 2000转染 C17.2神经干细胞,挑选稳定表达的克隆,用 RT-PCR及 Western Blot印迹分析鉴定。结果 pcDNA3.1-hygro-NTN质粒转染C17.2神经干细胞后有5个稳定表达的克隆生长,克隆1有NTN蛋白的高表达。结论 获得了稳定表达NTN蛋白的C17.2神经干细胞克隆,为移植治疗PD打下了坚实的实验基础。

Objective To explore the neuroprotection through delivery of Neurturin by C17.2 neural stem cell in a mouse of Parkinson s disease and provide with gene- modified neural stem cell. Methods We cloned the mouse prepro-neurturin by RT-PCR, subcloned it into pBluescriptII SK( + ), and identified the right clone by digestion of restriction enzyme and sequencing. Then we stably transfected pcDNA3 .1-hygro-NTN into C17.2 neural stem cells with lipofectamine 2000, and identified the protein expression of NTN gene by RT-PCR and Western Blot. Results We got five Neurturin clones after stably transfecting pcDNA3. 1-hygro-NTN, of which clone 1 overexpressed NTN protein. Conclusion We got Neurturin clones stably overexpressed the transgene, which made solid experiment basis for the treatment of PD.