枇杷EjAO基因的克隆与表达分析

发布时间：2018-07-21 15:41

【摘要】：果实的成熟和衰老是个复杂的生物学过程,一般认为活性氧(ROS)在其中起着重要作用。本研究采用cDNA末端快速扩增技术(RACE)和RT-PCR相结合的方法,从白肉枇杷成熟期果实中成功克隆到一个编码抗坏血酸氧化酶(AO)的基因,命名为EjAO。该基因全长2 174 bp,包含了长度为1 605 bp的开放阅读框。该阅读框编码的氨基酸数目为534个,所形成的蛋白质预测分子量为59.39 kD。实时荧光定量PCR结果显示EjAO在根和果实中高丰度表达,并且在白肉枇杷成熟期果实中的表达量远远高于红肉枇杷。核苷酸多态性分析显示EjAO基因在红肉枇杷和白肉枇杷共有五处多态性位点,其中T719C和A730C均导致氨基酸发生改变,推测可能对EjAO的酶学活性产生影响。EjAO的克隆和多态性位点的鉴定为阐明枇杷果实成熟与衰老机制及枇杷遗传改良育种提供一定的理论依据。
[Abstract]:Fruit maturation and senescence is a complex biological process in which reactive oxygen species (Ros) play an important role. In this study, a gene encoding ascorbic acid oxidase (AO) was successfully cloned from the mature fruit of loquat by rapid amplification of cDNA terminal (race) and RT-PCR, named EjAO. The gene is 2 174 BP in length and contains an open reading frame of 1 605 BP in length. The number of amino acids encoded by the reading frame is 534, and the predicted molecular weight of the protein is 59.39 KD. The results of real-time fluorescence quantitative PCR showed that EjAO was highly abundant in root and fruit, and the expression level in white flesh loquat fruit was much higher than that in red loquat fruit. Nucleotide polymorphism analysis showed that EjAO gene had five polymorphic loci in red loquat and white loquat, among which T719C and A730C caused amino acid changes. It is speculated that the enzyme activity of EjAO may be affected. The cloning and polymorphic site identification of EjAO may provide a theoretical basis for elucidating the mechanism of mature and senescence of loquat fruit and for genetic improvement and breeding of loquat.
【作者单位】：四川农业大学园艺学院;四川省农业科学院园艺研究所农业部西南地区园艺作物生物学与种质创制重点实验室;
【基金】：四川省财政厅创新能力提升工程重点实验室专项(2013JCYJ-004);四川省财政厅现代农业技术创新与示范专项资金项目(2014CXSF-015) 农业部重点实验室补助项目(2015JSCX-036)共同资助
【分类号】：Q943.2;S667.3

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