靶向cofilin2基因RNAi对鸭病毒性肠炎病毒增殖的影响

Effect of RNAi Targeting cofilin2 Gene on Duck Enteritis Virus Proliferation

旨在探讨cofilin2基因的表达在鸭病毒性肠炎病毒(DEV)增殖过程中的影响。本研究通过已获得的鸭源cofilin2全基因序列(GenBank登录号:MF434775),设计出4对shRNA序列,插入载体中,并转染到鸭胚成纤维细胞中,经FQ-PCR方法和荧光显微镜观察筛选出具有较高干扰效率的shRNA序列,用干扰效率较高的shRNA序列转染到鸭胚成纤维细胞中抑制cofilin2基因的表达,再检测cofilin2在DEV复制过程中表达情况。结果显示:4对shRNA干扰质粒经转染至鸭胚成纤维细胞后,经荧光显微镜观察,构建的4对shRNA干扰质粒均成功转染并得到表达;4对干扰质粒沉默率分别为4.96%±0.78%、41.12%±2.94%、17.55%±2.07%、54.02%±1.73%,以cofilin2-86的沉默效率最高;cofilin2-86转染到鸭胚成纤维细胞,经DEV感染不同时间,其DEV核酸表达量明显下降。综上所述,靶向cofilin2基因RNA干扰在DEV复制和增殖过程发挥重要作用。

This study aimed to investigate the effect of cofilin2 gene expression on the proliferation of duck enteritis virus(DEV).The complete gene sequence of cofilin2 from duck(GenBank:MF434775),four pairs of shRNA sequences were designed,inserted into vectors and transfected into duck embryo fibroblasts.shRNA sequences with high interference efficiency were screened by FQ-PCR and fluorescence microscope observation.The shRNA sequences with high interference efficiency were transfected into duck embryo fibroblasts to inhibit cofilin2 gene expression.Then check how cofilin2 is expressed during DEV replication.Result were as follows:Four pairs of shRNA interfering plasmids were successfully transfected into duck embryo fibroblasts and expressed by fluorescence microscopy.The silences were 4.96%±0.78%,41.12%±2.94%,17.55%±2.07%,54.02%±1.73%,respectively,and cofilin2-86 was the most efficient.After cofilin2-86 transfection into duck embryo fibroblasts and DEV infection,the expression of DEV nucleic acid decreased significantly at different time.In conclusion,RNA interference targeting cofilin2 gene plays an important role in DEV replication and proliferation.