蜻蜓凤梨AfACO1基因的克隆及乙烯响应特性分析

Cloning and Expression Analysis of AfACO1 Response to Exogenous Ethephon Treatment in Aechmea fasciata

观赏凤梨是一种重要的热带花卉。生产上常人工施加外源乙烯或乙烯衍生物促进凤梨科植物开花,但作用的分子机制不清楚。以热带观赏花卉蜻蜓凤梨(Aechmea fasciata)为材料,利用转录组数据结合cDNA末端快速扩增(rapid amplification of cDNA ends,RACE)技术克隆了乙烯生物合成过程中限速酶1-氨基环丙烷-1-羧酸(1-aminocyclopropane-1carboxylate,ACC)氧化酶(ACC oxidase,ACO)的一个编码基因,将其命名为AfACO1。AfACO1cDNA全长732bp,编码156个氨基酸。理化性质分析表明,该蛋白分子质量为17.47ku,等电点为8.46。AfACO1蛋白有保守的抗坏血酸和辅因子亚铁离子结合位点。进化树分析结果表明,AfACO1与水稻的2个OsACO1-likes蛋白亲缘关系最近,且进一步的结构域分析表明,它们拥有相似的保守基序。实时荧光定量PCR(quantitative real time PCR,qPCR)结果表明,AfACO1在抽薹39d后的成株心叶中表达量最高,且施加外源乙烯利后,AfACO1的表达量逐渐上调。结果表明AfACO1可能是蜻蜓凤梨成熟期乙烯生物合成的关键酶之一。研究结果为解析外源乙烯调控凤梨科植物开花的机制奠定了基础。

Ornamental bromeliads are important tropical flowering plant.Ethylene or substances that could release ethylene(i.e.ethephon,acetylene,etc)are widely used to promote flowering of bromeliads,however,the molecular mechanism is still unclear.With the ornamental bromeliads Aechmea fasciata as material,based on its transcriptome data and rapid amplification of cDNA ends(RACE)technology,a1-aminocyclopropane-1carboxylate(ACC)oxidase(ACO)gene named as AfACO1which might encode one of the rate-limiting enzyme was identified during ethylene biosynthesis.The length of AfACO1cDNA was732bp,which could encode a156-amino acids deduced protein.The molecular weight of the protein was17.47ku,with a theoretical pI of8.46.AfACO1contained both conserved ascorbic acid and cofactor Fe^2+binding sites.Phylogenetic analysis indicated that AfACO1was close to two OsACO1-likes.Furthermore,MEME motif analysis showed that it had similar conserved motifs.Quantitative real time PCR(qPCR)results showed that AfACO1had highest expression level in central leaves of39-day-after-flowering(DAF)adult plants.In addition,the expression of AfACO1transcripts in central leaves of adult plants increased gradually before flowering after it was treated with exogenous ethephon.All the results above indicated that AfACO1might be at least one of the rate-limiting enzyme during ethylene biosynthesis in Aechmea fasciata,which could provide a theoretical basis for flowering mechnism of bromeliads in the near future.