CDK5基因修饰神经干细胞体系的建立和分化特性的研究

目的建立携带CDK5与绿色荧光蛋白（GFP）的神经干细胞体系并观察CDK5对神经干细胞的作用。方法采用PCR、分子克隆与测序等技术成功构建CDK5-pEGFP表达质粒;并通过基因转染等技术将其转染入体外培养的神经干细胞中。结果（1）CDK5-pEGFP表达质粒经酶切、测序鉴定重组成功。（2）转染后神经干细胞分化24h后观察到表达CDK5的GFP荧光细胞已显示清楚的短突起,而对照组细胞的突起不明显;72h后表达CDK5的GFP细胞突起增长尤为明显,与对照组相比差异有统计学意义（P〈0.01）;GFP阳性细胞体积也增大,突起明显增多、加粗。结论CDK5的表达可促进分化后神经细胞突起的生长和延长,并能加快神经细胞的形态成熟。

Study on roles of CDK5 in neuronal differentiation by using system of genetically modified neural stem cell

Objective To investigate the possible roles of CDK5 in differentiation of neural stem cells.Methods The expression vector of CDK5-pEGFP was constructed by PCR and molecular cloning methods and identified by sequencing analysis.Then CDK5-pEGFP was transfected into the neural stem cell in vitro by using cell culture and gene transfection methods.Results(1) The expression vector of CDK5-pEGFP was identified success fully by sequencing analysis.(2) It was observed that the differentiated neural cells modified by CDK5-EGFP showing short processes after 24h.The differentiated GFP-positive cells revealed marked longer neurites than that of the control(transfected EGFP plasmid,P<0.01) after 72h,usually with enlarged soma and numerous number of processes.Conclusion The CDK5 expression can promote neurite outgrowth and extension.Therefore,it may play a role in the morphologic mature of the differentiated neural cell.