黄牡丹PlWDR3和PlWDR18转录因子基因的克隆与表达

Isolation and Expression of PlWDR3 and PlWDR18 Transcription Factor Genes in Paeonia lutea

以云南野生黄牡丹为试验材料,根据已构建的云南野生黄牡丹花瓣转录组数据库提供的WDR40蛋白的Unigene筛选得到19个与花色素合成相关的WDR40蛋白同源性高的Unigene序列,命名为PlWDR1~19.通过氨基酸序列比较和系统进化树分析,发现PlWDR3和PlWDR18可能参与调控黄牡丹的花色素合成。PlWDR3的ORF包含1个1 032bp的开放阅读框,编码1个344个氨基酸的蛋白,与苹果MdTTG1的亲缘关系最近,相似性达83.29%;PlWDR18的ORF包含1个1 035bp的开放阅读框,编码1个345个氨基酸的蛋白,与葡萄VvWDR2相似性最高,达93.77%。相对荧光定量PCR分析表明,PlWDR3在黄牡丹和紫牡丹的不同时期的表达模式基本相似,在黄牡丹和紫牡丹圆桃期表达量最高,在初开期表达量最低;PlWDR18在黄牡丹花发育初期呈上升趋势,在透色期达到最高峰,然后下降,在初开期达到最小值;在紫牡丹花发育初期亦呈上升趋势,在圆桃期达到最大值,然后逐渐下降,在盛开期降到最小值。PlWDR18在黄牡丹和紫牡丹花的各个发育时期的表达量均高于PlWDR3的表达量。推测PlWDR3和PlWDR18参与黄牡丹花色形成的调控,为今后深入探讨黄牡丹花色形成机制奠定基础。

In this study,19 unigene sequences that shared high homology with WDR40 transcription factor protein involved in plant anthocyanin biosynthesis were obtained from previous-constructed petal transcriptome database of Paeonia lutea and named PlWDR1-19.PlWDR3 and PlWDR18 were considered to be related to regulate anthocyanin biosynthesis by comparing analysis of amino acid sequence and phylogenetic tree analysis.And PlWDR3 contained a 1 032 bp ORF encoding 344 amino acid residues and PlWDR18 contained a 1 035 bp ORF encoding 345 amino acid residues,which both contained the typical WD40 structural domain.The predicted protein sequence of PlWDR3 shared high similarity with MdTTG1 （83.29%）,and PlWDR18 shared 93.77% similarity with VvWDR2.Relative real-time PCR analysis indicated that PlWDR3 showed similar expression pattern in different periods of P.lutea and P.delavayi,and reached the highest abundance at stage 2 while it reached the lowest level at stage 3.And PlWDR18 rose at the beginning stages of P.lutea with the peak at stage 3 and then declined while it had the highest expression level at stage 2 and then declined with the lowest level at stage 5.The results also showed that PlWDR18 expressed the higher abundance than PlWDR3 in all samples.In conclusion,we inferred that PlWDR3 and PlWDR18 might associate with the regulatory of anthocyanin biosynthesis in P.lutea and this would provide the basis for insight into the molecular mechanisms underlying tree peony yellow flower pigmentation.