黄牡丹花瓣转录组分析及不同开放阶段差异基因筛选

Transcriptome Analysis of Paeonia lutea and Its Differentially Expressed Genes Screening at Different Flower Opening Stages

为揭示黄牡丹花色形成过程中类黄酮生物合成相关基因的表达情况,本研究以云南野生黄牡丹为材料,采用Solexa测序技术对不同开放阶段的花瓣总RNA等量混合的样品进行转录组测序,经组装共得到88 330个Unigene,平均长度为603 bp。在此基础上,对“硬蕾期”、“初开期”和“盛开期”的花瓣建立差异表达谱,共获得7 790个表达差异基因。挑选GO功能显著性富集和Pathway显著性富集分析中富集在类黄酮生物合成代谢通路的差异基因Pl-CHS1、Pl-CHI1、Pl-THC2’GT和Pl-5GT1进行qRT-PCR表达模式验证。结果表明,随着花朵的开放和着色,Pl-CHS1的表达量逐渐上升;而Pl-CHI1和Pl-5GT1的表达量迅速降低;与“硬蕾期”相比,Pl-THC2’GT的表达量在“初开期”迅速上升,并且其高水平表达持续保持到花朵盛开。该结果与表达谱分析结果相一致。本研究为深入开展黄牡丹花色形成相关基因的克隆和功能验证提供了丰富的信息,并为揭示牡丹黄色花形成分子机制提供了帮助。

To elucidate the expression characteristics of flavonoid biosynthesis related genes during flower coloration, Paeonia lutea wild population in Yunan was taken as the material in this research, and the mixture samples of equal amounts of RNA from petals of different flower opening stages was constructed and sequenced using the Solexa/Illumina RNA-seq technologies. A total of 88 330 Unigenes were obtained by de novo assembly, with an average length of 603 bp. Based on the sequences of transcriptome, the digital gene expression profiles (DGE) among petals of unopened flower buds, initially opened flowers and fully opened flowers were established by using Solex technique, and 7 790 differentially expressed genes were obtained. Subsequently, the expression patterns of Pl-CHS1, Pl-CHI1, Pl-THC2'GT and Pl-5GT1, which were enriched in the flower pigmentation related pathway by gene ontology (GO) functional enrichment and pathway enrichment analyses, were further selected to be evaluated by qRT-PCR. The results indicated that the expression level of Pl-CHS1 increased gradually during flower opening and pigmentation;while mRNA levels of Pl-CHI1 and Pl-5GT1 decreased rapidly. Compared with the expression in petals of tight bud, Pl-THC2'GT expression level increased sharply in initially opened flower petals and remained high level until flower fully opened. The result is consistent with that of expression profile analysis. The Unigene sequences obtained in this study would provide cloning and functional verification of pigmentation-related genes in P. lutea, and be helpful for insight into the molecular mechanisms underlying tree peony yellow flower pigmentation.