Abstract: Using the resistant inbred line T32 and two susceptible inbred lines HuangC and Jiao 51 as experimental materials，we performed treatments with and without inoculation of Sporisorium reilianum.Samples were collected for high‑throughput sequencing（RNA‑seq） to screen differentially expressed genes（DEGs）. Based on the FPKM expression matrix of all samples，weighted gene co‑expression network analysis（WGCNA） was conducted after data processing to construct a weighted gene co‑expression network and identify modules significantly positively correlated with the disease resistance phenotype. Module‑trait cross hub candidate genes were screened by both module membership and gene significance，and then were intersected with the identified DEGs. The overlapping genes were subjected to KEGG pathway and GO term enrichment analysis. Combined with literature reviews，final disease resistance candidate genes were selected，to provide a theoretical basis for molecular disease resistance breeding.The results showed that high‑quality 154 Gb data were obtained by RNA‑seq，experimental reproducibility was good，and data was reliable. Differential expression analysis of genes revealed 10 563 DEGs（4 822 up‑regulated genes，5 741 down‑regulated genes）in T32 inoculation vs HuangC inoculation group（YTA vs YHA），10 808 DEGs（5 698 up‑regulated genes，5 110 down‑regulated genes）were selected in T32 inoculation vs Jiao 51 inoculation group（YTA vs YJA），and 1 335 DEGs（588up‑regulated genes，747 down‑regulated genes）in T32 inoculation vs T32 non‑inoculation group（YTB vs YTA），respectively.KEGG pathway enrichment analysis indicated that these DEGs were significantly enriched in carbon metabolism，biosynthesis of amino acids，phenylpropanoid biosynthesis，and MAPK signaling‑plant pathway and so on. WGCNA showed that two modules（brown and turquoise）were positively correlated with disease resistance，and 2 787 module‑trait cross hub candidate genes were screened.Among these，906 genes were enriched in KEGG pathways，primarily involved in carbon metabolism，biosynthesis of amino acids，cysteine and methionine metabolism. Thirty‑five intersection genes of module‑trait hub candidate genes with DEGs were obtained，16 of which obtained annotations in the NCBI database. KEGG pathway analysis of the above 35 genes showed that eight genes were enriched in pathways like metabolic pathway and plant hormone signal transduction，GO analysis revealed that six genes were enriched in protein dimerization activity，including five genes encoding transcription factors（three MADS，one bZIP and one bHLH ） and one gene encoding auxin‑responsive protein IAA24.Finally，ten disease resistance candidate genes were comprehensively selected，including two directly related to disease resistance［LOC100283065 and LOC100280308（TLP1）］，three genes involved in auxin and flavonoid metabolism［LOC100284018（CHI），LOC100273383 and LOC100280600（IAA24）］，and five genes encoding transcription factors（three MADS，one bZIP and one bHLH）］.

Key words: Maize, Head smut, RNA-seq, WGCNA, Candidate gene resistant to disease

摘要： 以玉米抗丝黑穗病自交系T32和2个感病自交系黄C、交51为试验材料，分别进行丝黑穗病菌接种与不接种处理，取样进行转录组高通量测序（RNA-seq），筛选差异表达基因。基于所有样本的FPKM表达矩阵，经处理后进行加权基因共表达网络分析（WGCNA），构建加权基因共表达网络，识别与抗病表型显著正相关的模块。以模块成员度和基因显著性共同筛选模块-性状交叉枢纽候选基因，并与差异表达基因取交集。对交集基因进行KEGG通路和GO条目富集分析，结合文献报道筛选最终的抗病候选基因，为抗病分子育种提供理论依据。结果表明，通过RNA-seq共获得154 Gb高质量数据，试验重复性良好、数据可靠。基因差异表达分析显示，T32接菌与黄C 接菌对比组（YTA vs YHA）中共筛选到10 563个差异表达基因（4 822个基因上调表达，5 741个基因下调表达），T32与交51接菌对比组（YTAvs YJA）中共筛选到10 808个差异表达基因（5 698个基因上调表达，5 110个基因下调表达），T32接菌与不接菌对比组（YTB vs YTA）中共筛选到1 335个差异表达基因（588个基因上调表达，747个基因下调表达）。KEGG富集分析发现，这些差异表达基因显著富集于碳代谢、氨基酸生物合成、苯丙烷生物合成和植物MAPK信号通路等通路。WGCNA分析鉴定出棕色和蓝绿色2个与抗病组正相关的共表达模块，共筛选到2 787个模块-性状交叉枢纽候选基因，其中906个基因富集到KEGG通路，主要涉及碳代谢、氨基酸生物合成、半胱氨酸和蛋氨酸代谢等。模块-性状交叉枢纽候选基因与差异表达基因交集获得35个基因，16个被NCBI注释。对35个基因进行KEGG通路富集分析发现，8个基因富集到代谢通路、植物激素信号转导等通路；进行GO条目富集分析发现，6个基因富集于蛋白质二聚化活性，包括5个转录因子基因（3个MADS、1个bZIP、1个bHLH）和1个生长素响应蛋白基因IAA24。最终综合筛选出10个抗病候选基因，包括直接抗病相关类2个［LOC100283065和LOC100280308（TLP1）］、生长素与类黄酮代谢类3个［LOC100284018（CHI）、LOC100273383 和LOC100280600（IAA24）］、转录因子类5 个［3 个MADS（LOC542257、LOC542040、LOC541765）、1个bZIP（LOC100191786）、1个bHLH（LOC100284818）］。

关键词: 玉米, 丝黑穗病, RNA-seq, WGCNA, 抗病候选基因