新型呼肠病毒S1基因与宿主细胞相互作用的初步研究

Preliminary Study on the Interaction between S1 Gene of a New Type Reovirus and Its Cell Receptor

目的筛选出新型呼肠病毒S1基因和宿主相互作用的关键区段,为后续的宿主受体蛋白筛选工作提供可靠的基础。方法将编码全长S1基因,以及N端和C端阅读框分别克隆入真核表达载体pIRSE2-EGFP,转染敏感细胞株鼠成纤维细胞(L929)和人宫颈癌细胞(Hela)后,通过荧光报告基因EGFP的表达分析,筛选出S1基因与宿主细胞相互作用时起决定性影响的区段。结果同等量的3种真核表达质粒在单独转染L929时,pGSN质粒转染后荧光表达量最大,pGSC质粒转染后荧光表达量最小。不同比例混合的pGS与pGSN进行共转染时,pGSN在转染质粒中的比例越高,荧光表达量也越高。在Hela细胞中的转染结果与L929相同。结论新型呼肠病毒R4株的S1基因N端阅读框编码区(62—406 bp)在S1基因转染时起关键作用。

Objective To study the key fragment which was important for the interaction between S1 gene of a new type reovirus and its cell receptor,and provide the basis for receptor screening.Method Three fragments,which coding full-length S1 and N and C terminal of S1 gene were cloned to pIRSE2-EGFP vector to get recombinant plasmids pGS,pGSN and pGSC.L929 and Hela cells were transfected with three recombinant plasmids,respectively,and the fragment which was involved for interaction of S1 gene and receptor was indicated by analysis the expression of reporter gene EGFP.Result When transfecting with equal amount of three recombinant plasmids,pGSN-treatment group showed the highest level of EGFP expression and pGSC-treatment group produced the lowest EGFP expression.Moreover,when co-transfecting with pGS and pGSN at different ratio,the higher ratio of pGSN the admixture comtained,thehigher level of EGFP expression could generate.Conclusion N terminal of S1 gene （62-406 bp） was essential for interaction between S1 gene and its cell receptor.