人源核不均一核糖核蛋白E1真核表达载体在神经细胞中的表达

摘要

背景：人源核不均一核糖核蛋白E1功能广泛，可参与神经系统骨架蛋白的表达。目的：为深入研究其在神经细胞中的作用，构建其真核表达载体，观察其在神经细胞中的表达。方法：利用真核表达载体pcDNATM4/His C，通过亚克隆构建核不均一核糖核蛋白E1-pcDNATM 4/His C重组质粒，经酶切、测序鉴定，通过转染神经细胞SH-SY5Y，采用western-blot，RT-PCR鉴定核不均一核糖核蛋白E1重组质粒的表达，并观察转染细胞的生长现象。结果与结论：成功构建了核不均一核糖核蛋白 E1的真核表达载体，mRNA 和蛋白水平上均证实了该质粒可在神经细胞SH-SY5Y中正确表达。SH-SY5Y细胞在转染核不均一核糖核蛋白E1后表现为加速生长。提示该蛋白对神经细胞的生长发育具有重要的作用。该载体为进一步研究核不均一核糖核蛋白E1在神经系统中的功能提供了前提条件。%BACKGROUND:The functions of homo heterogeneous ribonucleoprotein E1 are very wide. It can participate in the expression of skeleton proteins in the nervous system. OBJECTIVE:To construct the recombinant plasmid of homo heterogeneous ribonucleoprotein E1 and observe its expression in nerve cells for further studying the functions of it in neurocytes. METHODS:Using pcDNATM4/His C, the homo heterogeneous ribonucleoprotein E1 was subcloned into recombinant plasmid E1-pcDNATM 4/His C, fol owed by enzyming and sequencing. After SH-SY5Y cells were transfected with the recombinant plasmid, western blot analysis and real time RT-PCR were used to detect the expression of homo heterogeneous ribonucleoprotein E1 in SH-SY5Y cells. And the growth of SH-SY5Y cells was observed. RESULTS AND CONCLUSION:We successful y constructed the eukaryotic expressed vector of homo heterogeneous ribonucleoprotein E1. The recombinant plasmids were verified to express in SH-SY5Y cells correctly at mRNA and protein levels. And SH-SY5Y cells generated quickly after homo heterogeneous ribonucleoprotein E1 was over-expressed. The homo heterogeneous ribonucleoprotein E1 is an important protein in neural development. And this vector offers the premise for further studying its function in nervous system.