干旱胁迫下两个玉米自交系雄穗花器官分化期的转录组分析

Transcriptome Analysis of Floral Organ Differentiation Stage of Two Maize Inbred Lines under Drought Stress

【目的】运用现代分子生物学及高通量测序技术,在全基因组水平鉴定耐旱相关的基因,研究玉米雄穗花器官分化期对干旱胁迫的响应机理,为抗旱育种将提供新的理论基础。【方法】以干旱胁迫15 d和正常浇水的耐旱自交系“PHBA6”和干旱敏感自交系“吉63”花器官分化期的雄穗进行Illumina HiSeq 2000高通量测序分析,将测序得到的Unigene与玉米参考基因组进行比对,利用RPKM来衡量样本间的基因表达丰度,以|log2foldchange|≥1,P<0.05,FDR≤0.001筛选出样本间的差异表达基因,并通过Gene Onto 1ogy(GO)数据库、KEGG pathway数据库对筛选出的差异表达基因的功能和参与的调控路径进行分析。【结果】与正常水分处理相比,耐旱自交“PHBA6”和干旱敏感自交系“吉63”在干旱胁迫下分别有1394个和1300个显著差异表达基因,其中共有差异表达基因831个,包括上下调表达趋势是一致的822个基因和9个具有相反的表达模式基因。除这些基因外,各自分别还有563个和469个基因型特异的干旱响应基因,这些特异的干旱响应基因的G0富集分析的条目并不完全相同,除共同富集在刺激响应,细胞壁改变外,耐旱自交“PHBA6”还富集在激素合成调控,机氮化合物代谢、蛋白结合上,特异的干旱响应基因的KEGG显著性富集分析在2个自交系中也不完全相同,耐旱自交“PHBA6”还有部分差异基因富集在生物素代谢通路上,另外,筛选出耐旱自交“PHBA6”中在干旱胁迫下差异表达的3个脱水素基因和3个特有差异表达的bZIP、MYB和ERF转录因子。【结论】胁迫相关基因的特异性响应可能是材料间耐旱性存在差异的主要原因,耐旱自交“PHBA6”中的3个脱水素基因和3个特有差异表达的bZIP、MYB和ERF转录因子基因在玉米耐旱育种中可能有重要的利用价值。

【Objective】In order to analyze the mechanism of maize tassel to drought stress in the differentiation stage of male flower organs of maize,the identification of drought tolerance related genes at the whole genome level through modern molecular biology and high-throughput sequencing technology will provide a new theoretical basis for drought resistance maize varieties.【Method】High-throughput transcriptome sequencing by Illumina HiSeq 2000 was conducted using the tassels of drought-tolerant inbred line"PHBA6"and drought-sensitive inbred line"Ji 63"under flower organ differentiation stage to drought stress and normal irrigation for 15 days.The obtained Unigenes were compared with the maize reference genome,and the abundance of gene expression among different samples measured according to transcriptome data by RPKM,and the differentially expressed genes between samples screened by the standard of|log2foldchange|≥1,P<0.05,FDR≤0.001.Functions and pathway of those differentially expressed genes were also investigated using Gene Onto1ogy(GO)database and KEGG pathway database.【Result】Compared with samples under normal water treatment,1,394 and 1,300 significantly differentially expressed genes were detected respectively in drought-tolerant inbred“PHBA6”and drought-sensitive inbred line“Ji 63”,of which 831 differentially expressed genes included 822 genes with the same expression patterns and 9 genes with opposite.In addition to these genes,563 and 469 genotype-specific drought response genes were detected respectively in the two lines.The items of G0 enrichment for these specific drought response genes not exactly the same.Except for the common items of G0 enriched in stimulus response and the cell wall changing,the drought-tolerant inbred"PHBA6"had some items of G0 enriched in the regulation of hormone synthesis,metabolism of organic nitrogen compounds,and protein binding.The KEGG enrichment analysis illustrated that these pathways of specific drought-responsive genes not exactly the same in the two inbred lines.There some differential genes enriched in the biotin metabolism pathway in drought-tolerant inbred“PHBA6”.In addition,three dehydrin genes and three bZIP,MYB and ERF transcription factors screened differentially expressed in drought-tolerant inbred"PHBA6"under drought stress.【Conclusion】These specific drought-responsive genes the main reason for drought tolerance difference between the two 3 dehydrin genes and three bZIP,MYB and ERF transcription factors which were screened in drought-tolerant inbred"PHBA6"may have important utilization value in the breeding of drought resistance maize varieties.