辣椒苗期抗感疫病比较转录组学分析

Comparative Transcriptome Analysis of Phytophthora blight Resistance of Pepper at Seedling Stage

为了研究辣椒抗疫病分子机制,探究其与抗疫病相关的功能基因,通过对114份辣椒自然群体抗疫病鉴定,选取1份抗疫病辣椒材料和3份感疫病辣椒材料,利用IlluminaRNA-seq测序平台进行高通量转录组测序,将所得高质量的序列(Cleanreads)与Pepper_Zunla_1_Ref_v1.0参考基因组进行比对。结果显示,有78.95%~85.11%的Clean数据比对到唯一的基因组位点。采用RPKM法计算基因表达量,并在此基础上以FDR≤0.001和|log2Ratio|≥1为条件筛选出2组样本间差异表达基因。通过排列前10的GeneOntology(GO)分类可以看出,主要功能有氧化还原酶活性、碳氧裂解酶活性、过氧化物酶活性、代谢过程和逆境响应等过程。其中,有117个差异基因能归入KEGG通路,包括25个上调差异基因,92个下调差异基因。在这些通路中,包括泛素介导的蛋白水解作用、氧化磷酸化、植物激素信号转导、磷脂酶D信号通路、磷脂酰肌醇信号系统等过程。通过研究发现,辣椒抗疫病是一个高度复杂的过程,它由多个交叉通路调节,包括新陈代谢过程、防御反应、激素调节等,这为后期深入研究辣椒抗疫病分子机理奠定基础。

The objective of this study was to study the molecular mechanism of pepper phytophthora blight and to explore the functional genes related to phytophthora blight resistance. Based on the identification of phytophthora blight resistance in114 natural populations of pepper,one resistant material and three susceptible materials were selected as materials to perform high- throughput transcriptome sequencing by means of Illumina RNA-seq sequencing platform. The clean reads were selected to map the reference sequences of Pepper_Zunla_1_Ref_v1.0. The results showed that 78.95%-85.11% clean reads could be matched to unique genome locus. The gene expression level was calculated using RPKM method. The false discovery rate≤0.001 and the absolute value of |log 2 Ratio|≥1 were used as the threshold to judge the significance of gene expression differences. Finally,the functions and pathways of differentially expressed genes(DEGs)were annotated by comparing them with the Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG). According to the top10 GO classification,the main functions included the oxidoreductase activity,carboxylolyase activity,peroxidase activity,metabolic process and adverse reaction etc. Among them,117 DEGs could be classified into the KEGG pathway,including 25 up-regulated DEGs and 92 down-regulated DEG. These pathways were associated with the ubiquitin-mediated proteolysis,oxidative phosphorylation,plant hormone signaling,phospholipase D signaling,and phosphatidylinoinositol signaling systems. The disease resistance of pepper was mainly related to enzymatic activity,stress response,protein metabolism pathway,plant hormones and other regulations. It was found that the disease resistance of pepper was a highly complex process,which was composed of multiple cross channel regulation,including metabolic processes,defense responses and hormone regulation,etc. The results laid the foundation for further studying the molecular mechanism of pepper disease resistance.