羊传染性脓疱病毒感染山羊皮肤成纤维上皮细胞差异表达miRNA分析

Differential Expression Analysis of miRNA from Orf Virus Infected and Uninfected Goat Skin Fibroblast Cells

旨在研究羊传染性脓疱病毒(orf virus,orfv)感染山羊皮肤成纤维细胞(goat skin fibroblasts cell,GSF)对GSF细胞microRNA(miRNA)表达谱影响,探究miRNA在orfv感染过程中的作用及调控机制。分别提取感染和未感染orfv的GSF细胞总RNA,构建miRNA文库,利用高通量测序技术进行miRNA差异表达分析,对差异表达miRNA靶基因进行预测,并进行GO和KEGG分析,随机选取10个差异miRNA进行RT-qPCR验证。结果显示,orfv感染组和未感染GSF细胞组相比共有678个显著差异表达的miRNA(fold change≥1.5),其中,上调表达miRNA有509个,下调表达miRNA有169个,uniq_miRNA的Venn图分析显示,感染组和对照组共有的miRNA仅占8.21%;GO和KEGG分析显示,差异表达miRNA主要参与脂质代谢、受体及细胞因子信号转导等细胞生物学过程,RT-qPCR验证结果与高通量测序结果一致。本研究结果表明,orfv感染GSF细胞对其编码的miRNA有显著影响,获得大量GSF细胞编码的与orfv感染相关的差异miRNA,为进一步从宿主miRNA层面揭示orfv感染和致病机制提供了参考依据。

To study the effects of orfv infection on different miRNA expression profile in goat skin fibroblasts(GSF)cells,and understand the roles and regulatory mechanisms of miRNA during orfv infection,total RNA was isolated from GSF cells with or without orfv infection.Differential expressed miRNAs were obtained and analyzed with high-throughput sequencing technology after the miRNA library being constructed.The target genes of differential expressed miRNA were predicted and analyzed by GO and KEGG methods.Ten different miRNAs were randomly selected for RT-qPCR verification.Results indicated that there were total 678 differential expressed miRNAs(fold change≥1.5)between orfv infected and control group,509 were up-regulated miRNAs,169 were down-regulated miRNAs,and only 8.21%were conjunct miRNAs between infection group and control group according to the Venn diagram analysis of uniq_miRNA.GO and KEGG analysis showed that the differentially expressed miRNAs were mainly involved in cell biological processes such as lipid metabolism,receptor and cytokine signal transduction.RT-qPCR results of miRNA were consistent with high-throughput sequencing results.In a word,this study obtained a large number of different miRNAs encoded by GSF cells and it indicated that infection of GSF cells with orfv has a significant impact on the miRNA,which provide a reference for further revealing the infection and pathogenesis mechanisms of orfv at the host cells miRNA level.