Mob1a真核表达载体构建方法及对MEN通路基因表达的影响研究

Construction of Mob1a eukaryotic expression vector and its effect on MEN pathway gene expression

目的探讨Mobla真核表达载体构建方法及对MEN通路基因表达的影响。方法利用PCR扩增技术扩增出Mobla编码基因,将其构建到真核表达Mobla载体上,利用脂质体将构建的真核表达载体瞬时转染干细胞,采用Western blot检测Mobla蛋白是否表达,采用荧光素梅报告基因实验对MEN通路基因表达活性的影响。结果本研究成功构建Mobla真核载体,经过小鼠肝脏细胞转染后能对其蛋白质进行提取,Western blot显示:Mobla质粒在小鼠肝脏细胞中表达水平未见异常。荧光素酶报告基因试验进结果显示:Mobla能抑制MEN通路基因信号,能抑制TNF-a刺激引起的MEN通路转录活性。结论采用PCR扩增技术能成功扩增、构建出Mobla真核表达载体,并且对MEN通路基因表达存在一定的影响。

Objective To investigate the effects of Mobla eukaryotic expression vector construction method and the expression of MEN pathwaygenes.Methods PCR amplified Mobla gene encoding by PCR,cloned into eukaryotic expression vector Mobla,to construct the eukaryotic expressionvector transfected stem cells by liposome was detected by Mobla Western blot is the expression,activity of fluorescent reporter gene expressionon Al MEN pathway genes.Results This study successfully constructed Mobla eukaryotic vector after transfection in mouse liver cells can be carriedout after the protein Western extraction,blot showed that the expression level of Mobla plasmid showed no abnormalities in mouse liver.The luciferasereporter gene into test results showed that Mobla could inhibit MEN signaling pathway genes,can inhibit the TNF-a pathway induced bystimulation of MEN transcriptional activity.Conclusion Using PCR amplification technique can be successfully amplified,construct the eukaryotic expression vector Mobla and the expression of MEN pathway genes have a certain influence.