猕猴桃AcNCED的克隆与表达分析

Cloning and Expression Analysis AcNCEDs in Kiwifruit

【目的】9-顺式环氧类胡萝卜素双加氧酶(NCED)是类胡萝卜素降解酶,同时是脱落酸(ABA)合成的关键酶。为探明AcNCED基因家族各成员在猕猴桃果实发育期的表达特性。【方法】利用RT-PCR技术从猕猴桃中分离4个NCED基因,分别命名为AcNCED1-4,并用qRT-PCR技术分析它们在猕猴桃不同组织和果实发育期的表达。【结果】AcNCED1、AcNCED2、AcNCED3和AcNCED4的开放阅读框(ORF)分别为1 755、1 830、1 797和1 788 bp,它们分别编码585、610、599和596个氨基酸。经预测,AcNCED1、AcNCED3和AcNCED4蛋白定位于叶绿体,AcNCED2蛋白定位于细胞质,均无信号肽,属于非分泌蛋白。qRT-PCR分析表明,AcNCEDs在茎中表达量最高,根中最低,在果实发育前期及成熟期高表达。【结论】4个基因编码的蛋白都较为保守且表达量均具有组织特异性,为进一步探明AcNCED的生物学功能奠定基础。

【Objective】9-cis-epoxy carotenoid dioxygenase(NCED)is an enzyme involving in carot-enoid degrading and a key enzyme in the synthesis of abscisic acid(ABA).To explore the expression characteristics of all members of the AcNCED gene family during kiwifruit development.【Method】Four NCED genes were cloned from kiwifruit by RT-PCR,named AcNCED1-4,and their expression in vari-ousduring tissues and fruit developmental stages and in different tissues of both varieties was analyzed by qRT-PCR.【Result】The open reading frames(ORF)of AcNCED1,AcNCED2,AcNCED3 and AcNCED4 are 1755,1830,1797 and 1788 bp,respectively,which encode 585,610,599 and 596 amino acids,respectively.It was predicted that AcNCED1,AcNCED3 and AcNCED4 proteins are localized in chloro-plast,while AcNCED2 protein in cytoplasm,all of which lacking signal peptide and belong to non-secreted proteins.The qRT-PCR analysis showed that AcNCEDs has the highest expression level in the stem,the lowest in the roots,and high expression during early fruit development and maturity.【Conclusion】The proteins encoded by all four genes are highly conserved and exhibit tissue-specific expression,which lays the foundation for further exploring the biological function of AcNCED.