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蝴蝶兰“大辣椒”APETALA1基因的克隆及表达

Molecular Cloning and Expression of APETALA1 Gene From Phalaenopsis ‘Big Chili’

  • 摘要: 以蝴蝶兰"大辣椒"为试材,采用RACE技术克隆蝴蝶兰APETALA1(AP1)基因的cDNA全长,分析基本生物学信息,并初步探索其在花芽分化过程中的作用。结果表明:AP1基因的cDNA全长为1 155 bp,开放阅读框(open reading frame)752 bp,编码250个氨基酸,含有MADS盒与K盒等保守功能结构域。同源性比对结果显示,蝴蝶兰"大辣椒"与朵丽蝶兰、金蝶兰、文心兰、石斛兰和马兜铃等植物的MADS蛋白有较高的相似性,同源性均在80%以上;系统进化树分析显示,蝴蝶兰"大辣椒"AP1基因与朵丽蝶兰聚类关系最近;实时荧光定量PCR检测发现,AP1基因在叶片、根和花葶中均有表达,但表达量有差异。同一器官不同发育时期比较显示,叶片的AP1基因表达量没有明显的时空差异;根AP1基因的表达量随着花芽分化的进程呈递减趋势,其中始花期的表达量最低,盛花期最高;而花葶中AP1基因表达量随着花芽分化呈增加趋势,在盛花期表达量达到峰值。同一时期不同器官比较,花葶中AP1的表达量显著高于同一时期的叶片和根(P<0.05)。由此,推测AP1基因在调控蝴蝶兰花芽分化过程中发挥重要作用。

     

    Abstract: Taking Phalaenvpsis ’Big Chili’ as experiment material,cloning full length cDNA of Phalaenopsis APETALA1 gene by RACE and analysis of basic biological infamati.The results showed full-length cDNA of AP1 gene was 1 155 bp with a 752 bp open reading frame encoding a putative protein of 250 amino acids.It contained conservative functional domains such as MADS-box and K-box.The results of homology comparison showed that the MADS proein of Phalaenopsis ’Big Chili’ had high similarity with other plants,such as Dory butterfly,Cymbidium formosans,Oncidium,Dendrobium and Aristolochia,and the homology was more than 80%.Phylogenetic tree analysis showed that the AP1 gene of Phalaenopsis ’Big Chili’ had the closest relationship with the clustering of Dory butterfly.qRT-PCR showed that the mRNA of AP1 was widely expressed in all tissues tested(leaves,roots and pedicels),but the expression level was different.There was no signficant difference of AP1 gene expression in leaves of different developmental stages of the same organ.The expression level of AP1 gene in roots decreased with the process of flower bud differentiation,the lowest level at the beginning flowering stage,and the highest at the full flowering stage;while the expression of AP1 gene in the scape increased with the flower bud differentiation,and the highest at the full flowering stage.Comparison of different organs in the same period,the expression of AP1 in pedicel was significantly higher than that in leaves and roots(P<0.05).Therefore,we speculate that AP1 gene plays an important role in the regulation of flower bud differentiation in Phalaenopsis.

     

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