D3基因在抗病防卫反应中的转录调控研究

Effects of the D3 gene on transcriptional regulation and its role in defense responses

水稻白叶枯病(bacterial leaf blight,BLB)是由水稻黄单胞菌属水稻致病变种(Xanthomonas oryzae pv.oryzae,Xoo)引起的细菌性病害,严重影响水稻产量。随着基因技术的发展,编辑抗病基因是提高水稻抗病性降低该病害的发生的一种行之有效的方法。本研究首先对D3基因编辑株系和野生型进行白叶枯病菌接种实验来确定D3基因对于白叶枯病菌的抗性,其次利用高通量测序技术对已筛选出的遗传纯合D3基因编辑株系进行转录组分析,同时结合基因差异表达分析、基因家族分析和富集分析挖掘其在抗病相关通路上的潜在作用,并通过实时荧光定量PCR(qRT-PCR)技术对部分差异表达基因(differential expressed genes,DEGs)进行验证。结果表明:接种白叶枯病菌后D3基因被显著诱导,同时D3基因编辑株系均表现出不同程度的白叶枯病抗性。转录组数据显示,与野生型相比,在转录组水平共鉴定到8184个差异表达基因,包含4201个上调基因和3983个下调基因,3组均呈现差异表达的相关基因共有359个。GO功能富集分析表明,D3基因编辑株系与野生型在生物学途径、细胞组分和分子功能上的相关基因均呈显著差异表达。KEGG代谢途径富集分析发现,DEGs显著富集于植物与病原菌互作、植物激素信号转导以及苯丙烷生物合成途径。本研究丰富了水稻D3基因在抗病相关基因转录调控方面的信息,为D3基因抗病功能研究拓展了思路。

Bacterial leaf blight is a bacterial disease caused by Xanthomonas oryzae pv.oryzae(Xoo).Bacterial leaf blight can severely reduce rice yields.With the development of molecular biotechnology,editing potential disease resistance genes has become a method to improve rice disease resistance and reduce the loss of the disease.In this study,D3 gene edited lines and wild-type inoculation experiments were used to determine D3 gene resistance and high-throughput sequencing technology was employed to analyze the transcriptomes of three different D3 gene-edited lines,and the results showed that compared with the wild type,D3 gene was significantly induced,while the D3 edited lines showed different degrees of blight resistance and a total of 8184 DEGs were identified at the transcriptome level,including 4201 up-regulated genes and 3983 down-regulated genes,among which 359 DEGs were found simultaneously differentially expressed in the three groups.After that,this part of differentially expressed genes(DEGs)was verified by quantitative PCR.GO functional enrichment analysis showed that the genes related to biological pathways,cellular components and molecular functions were significantly differentially expressed in D3 gene-edited lines compared with the wild-type.KEGG pathway enrichment analysis found that DEGs were significantly enriched in plant-pathogen interaction,plant hormone signal transduction and phenylpropane biosynthesis pathways.Further experiments showed that,compared with the wild type,pathogen-related genes(PRs)in D3 gene-edited lines were significantly induced after inoculation with Xoo,and D3 gene-edited lines showed different degrees of bacterial blight resistance,thus further confirmed the role of D3 in bacterial blight resistance.This study enriches the role of the rice D3 gene in affecting transcriptional regulation,especially its impact on plant disease resistance-related genes.At the same time,this study expands the ideas for the research on the disease resistance mechanism of D3 gene.