番茄斑萎病毒侵染辣椒苗期的差异基因表达分析

Analysis of Differential Gene Expression in Pepper Seedlings Infected by Tomato spotted wilt orthotospovirus

为了研究番茄斑萎病毒与辣椒的互作机制,探究病毒侵染后mRNA表达谱,本研究选用‘萧新19’为试验材料,利用Illumina RNA-seq测序平台进行转录组测序,将所得高质量的序列(Clean reads)与CM334_Annuum.v.2.0参考基因组进行比对。结果显示,有74.09%~76.91%数据比对到唯一的基因组位点。采用FPKM法计算基因的表达量,并在此基础上以FDR≤0.05和|log_(2)FC|≥1为条件筛选出2组样本间差异表达基因。对差异基因进行Gene ontology(GO)和KEGG富集分析,通过排列前5的GO分析发现,主要分子功能有催化活性、结合等过程,差异表达基因显著富集到次生代谢物的生物合成、苯丙烷类生物合成、亚麻酸代谢等过程。从苯丙烷类合成途径和转录因子中挑选基因用RT-qPCR来验证转录组测序的真实性。通过研究发现,辣椒与番茄斑萎病毒互作是一个复杂的过程,它由多个交叉通路调节,包括苯丙烷类生物合成、类黄酮生物合成、新陈代谢过程等,这为后期深入研究辣椒与番茄斑萎病毒互作机理奠定基础。

In order to study the interaction mechanism between Tomato spotted wilt orthotospovirus and pepper,and explore the mRNA expression profile after virus infection,’Xiaoxin19’was used as pepper material,transcriptome sequencing using the Illumina RNA-seq sequencing platform,and the high-quality sequence(Clean reads)was compared with the CM334_Annuum.v.2.0 reference genome.The results showed that 74.09%~76.91%of the data were aligned to the unique genomic locus.The gene expression was calculated by FPKM method,and the differentially expressed genes between the two groups were screened on the condition of FDR≤0.05 and|log_(2)FC|≥1.Gene ontology(GO)and KEGG enrichment analysis of differential genes showed that the main molecular functions were catalytic activity and binding,and differentially expressed genes were significantly enriched in secondary metabolites biosynthesis,phenylpropane biosynthesis and linolenic acid metabolism by GO classification of the top5.RT-qPCR was used to verify the authenticity of transcriptome sequencing by selecting genes from phenylpropane synthetic pathways and transcription factors.Through the study,it is found that the interaction between pepper and Tomato spotted wilt orthotospovirus is a complicated process,which is regulated by several cross pathways,including phenylpropane biosynthesis,flavonoid biosynthesis,metabolic processes,etc..This lays the foundation for the in-depth study of the interaction mechanism of pepper and Tomato spotted wilt orthotospovirus.