敲除小RNA gcvB后沙门菌的转录组分析  

mRNA-Seq Whole-transcriptome Analysis of sRNA gcvB Deletion Background in Salmonella

作　　者：刘丽娟[1,2] 董然然 王开功[1,2] 程振涛 文明[1,2] 温贵兰 李晨[1,2] 杨琦[1,2] 周碧君[1,2] LIU Lijuan;DONG Ranran;WANG Kaigong;CHENG Zhentao;WEN Ming;WEN Guilan;LI Chen;YANG Qi;ZHOU Bijun(College of Animal Science,Guizhou University,Guiyang 550025,China;Institute of Animal Diseases,Guizhou University,Guiyang 550025,China)

机构地区：[1]贵州大学动物科学学院,贵阳550025 [2]贵州大学动物疫病研究所,贵阳550025

出　　处：《畜牧兽医学报》2019年第4期840-850,共11页ACTA VETERINARIA ET ZOOTECHNICA SINICA

基　　金：国家自然科学基金(31602065);贵州省科学技术基金计划项目(黔科合基础[2016]1047);贵州省科技创新人才团队建设项目(黔科合人才团队[2015] 4016)

摘　　要：为筛选与沙门菌小RNA gcvB相关的靶基因,采用高通量的转录组测序(RNA-seq)技术对野生型沙门菌和敲除小RNA gcvB沙门菌的mRNA进行对比分析,全面地预测GcvB的靶基因的数量与种类,通过GO和KEGG数据库对筛选基因进行比对注释,进一步分析筛选基因的主要功能及涉及的生物过程。并利用荧光定量PCR对部分差异表达基因进行验证。结果显示:差异表达基因共1 244个,其中基因表达上调678个,基因表达下调566个。GO富集分析显示,差异表达基因主要涉及氧化还原活性、铁离子结合、运动活性等分子功能,细菌鞭毛的细胞成分,细胞呼吸、钴胺素代谢过程和钴胺素生物合成过程等生物过程。KEGG数据库分析显示,差异表达基因主要参与细菌趋化性、卟啉和叶绿素代谢、不同环境中的微生物代谢、双组分系统、甲烷代谢等14个信号通路。对筛选出的10个差异表达基因进行荧光定量PCR验证,结果显示:差异表达基因相对表达量与RNA-seq测序结果基本一致。本研究为进一步探明小RNA gcvB与靶基因相互作用、小RNA的调控机制,以及沙门菌致病机制奠定了基础。In this article,high-throughput transcriptome sequencing(RNA-seq)technology was used to screen potential regulatory target genes of small RNA(sRNA)gcvB in Salmonella.Comparative analysis of mRNA in wild-type strain and knockout small RNA gcvB strain has been used.The number and type of GcvB target genes were comprehensively predicted,the main functions and the biological processes of the selected genes were compared by the GO and KEGG databases for further analysis.Some differentially expressed genes were verified by real-time PCR.The results showed that there were 1 244 differential expression genes have been screened out,including 678 up-regulated genes and 566 down-regulated genes.GO enrichment analysis showed that the differentially expressed genes mainly involved molecular functions such as redox activity,iron binding,locomotor activity,cellular components of bacterial flagella,cellular respiration,cobalamin metabolism,and cobalamin biosynthesis.KEGG database analysis showed that differential expression genes mainly participated in 14 signaling pathways,including bacterial chemotaxis,porphyrin and chlorophyll metabolism,microbial metabolism in different environments,two-component systems and methane metabolism.Ten differentially expressed genes in WT strain andΔGcvB strain were selected from transcriptome analysis and carried out the further Real-time PCR test,and the results show that the trend of gene expression is consistent in both transcriptome analysis and in Real-time PCR test.The study lays the foundation for further exploration of the interaction between sRNA GcvB and target genes,the regulation of sRNA and the pathogenic mechanism in Salmonella.

关 键 词：沙门菌 小RNAgcvB 转录组测序 靶基因 

分 类 号：S852.61[农业科学—基础兽医学]