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采用RT-PCR和RACE技术,从观赏向日葵新品种闽葵3号黄色花瓣中克隆获得类胡萝卜素合成途径关键酶基因ZDS,命名为Ha ZDS(Gen Bank No.KF263657),该基因c DNA全长2069 bp,具有一个1761 bp的完整开放阅读框(ORF),编码587个氨基酸。推导氨基酸序列分析表明,Ha ZDS蛋白在N-末端区域含有一个叶绿体转运肽序列,含有一个典型的氨基氧化酶结构域和NAD(P)结合结构域,存在一个特征序列。系统发育分析表明,观赏向日葵Ha ZDS与万寿菊、菊花蛋白的同源性分别达96.4%、90.1%,亲缘关系较近。利用实时荧光定量PCR技术分析结果显示,Ha ZDS基因在盛花期表达量最高,不同组织中的表达量舌状花瓣>绿色管状花>苞片>叶片>黑色管状花,随着花瓣黄色的加深,基因表达量逐渐增加。推测Ha ZDS基因在转录水平上参与观赏向日葵花色形成的调控。
The key enzyme gene ZDS of carotenoid biosynthesis pathway was cloned from ornamental sunflower new cultivar Mau Mau 3 by RT-PCR and RACE techniques, and named as Ha ZDS (Gen Bank No.KF263657) 2069 bp, with a 1761 bp complete open reading frame (ORF) encoding 587 amino acids. The deduced amino acid sequence analysis showed that the Ha ZDS protein contains a chloroplast transit peptide sequence in the N-terminal region, which contains a typical amino-oxidase domain and NAD (P) -binding domain, and has a characteristic sequence. Phylogenetic analysis showed that the homologies of ornamental sunflower Ha ZDS with marigold and chrysanthemum proteins were 96.4% and 90.1%, respectively, and their genetic relationship was close. The results of real-time fluorescence quantitative PCR showed that the expression of Ha ZDS gene was the highest at full flowering stage. The expression of Ha ZDS gene in different tissues was greater than that of tongue-shaped petals> green tubular flowers> bracts> leaves> black tubular flowers. Gene expression increased gradually. It is speculated that the Ha ZDS gene is involved in the regulation of sunflower flower formation at the transcriptional level.