鸭RIG-1基因的组织表达及体外抗禽流感病毒活性研究  

Detection of RIG-1 expression in duck tissues and its anti-avian influenza virus activity in vitro

作　　者：张雅春[1] 胡卫杰[1] 王建超[1] 王伟[1] 邓瑞坡[1] 李越[1] 赵颖慧[1] 孟庆文[1] 

机构地区：[1]中国农业科学院哈尔滨兽医研究所兽医生物技术国家重点实验室/实验动物研究室,黑龙江哈尔滨150001

出　　处：《中国预防兽医学报》2013年第12期955-959,共5页Chinese Journal of Preventive Veterinary Medicine

基　　金：国家自然基金项目(30771615);科技重大专项(2009ZX08006-01B)

摘　　要：视黄酸诱导基因蛋白1(RIG-1)是一类模式识别受体,在机体抗病毒天然免疫过程中发挥重要作用。为检测金定鸭RIG-1基因的组织特异性表达水平和体外抗禽流感病毒(AIV)活性,本研究采用RT-PCR方法扩增鸭RIG-1基因并克隆于pcDNA3.1(+)中构建重组真核表达质粒pcDNA-RIG-1,应用荧光定量PCR方法检测该基因在鸭不同组织中的特异性表达水平;并将pcDNA-RIG-1转染DF-1细胞,通过病毒TCID50测定、间接免疫荧光、荧光定量PCR的方法检测RIG-1的抗AIV活性。结果显示,从金定鸭脾脏中扩增的RIG-1基因的CDS序列大小为2 802 bp,编码934个氨基酸;RIG-1基因在不同组织中的表达各不相同,在鸭的脾脏和肾脏中表达量较高,肝脏中度表达,心脏、肺脏和肌肉低度表达;pcDNA-RIG-1转染DF-1细胞组与pcDNA3.1(+)转染组相比,病毒TCID50及相对表达量显著降低,IFN-β及Mx的相对表达量显著升高,表明RIG-1产生了显著的抗AIV作用。本研究为禽类RIG-1蛋白功能和家禽的天然免疫研究奠定了基础。Acting as a pattern recognition receptor, the retinoic acid inducible gene-1 （RIG-l） plays an important role in innate immunity for antivirus. To investigate the RIG-1 gene expression in different tissues and its antiviral activity in vitro, the duck RIG-1 gene was amplified by RT-PCR and cloned into pcDNA3.1 （＋） to construct pcDNA-RIG-1. The sequencing result showed that the duck RIG-1 gene was 2,802 bp encoding 934 amino acids. In addition, the expression of RIG-1 in different tissues was qualified by SYBR Green-based real time RT-PCR, the results showed that mRNA of RIG-1 expressed in a high level in kidney and spleen, and middle level in liver, but low level in heart, lung and muscle. Furthermore, the replication of avian influenza virus （A1V） was significantly inhibited in pcDNA-RIG-1 transfected DF-1 cells detected by standard titration method, indirect immunofluorescence assay and real time RT-PCR. Evidently, the results also displayed that the expressions of IFN-[3 and Mx were significantly increased in pcDNA-RIG-1 transfected DF-1 cells, resulting in inhibition of the avian influenza virus replication. These data demonstrated that duck RIG-1 has a significantly antiviral activity in vitro.

关 键 词：RIG-1 组织表达 抗病毒 

分 类 号：S852.65[农业科学—基础兽医学]