无核荔枝ABA生物合成关键酶LcNCED基因克隆及其在生理落果阶段中的表达分析  

Expression Analysis and Cloning of NCED Gene During Physiological Fruit Drop Period of Seedless Litchi Fruit

作　　者：陈哲[1] 胡福初[1] 年宇薇 吴凤芝[1] 范鸿雁[1] 陈业光[1] 王祥和[1] CHEN Zhe, HU Fuchu, NIAN Yuwei, WU Fengzhi, FANG Hongyan, CHEN Yeguang, WANG Xianghe(Institute of Tropical Fruit Trees, Halnan Academy of Agricultural Sciences/Hainan Provincial Key Laboratory of Tropical Fruit Tree Biology of Hainan /Haikou Investigation Station of Tropical Fruit Trees, Ministry of Agriculture, Haikou, Hainan 571100, Chin)

机构地区：[1]海南省农业科学院热带果树研究所海南省热带果树生物学重点实验室农业部海口热带果树科学观测实验站,海南海口571100

出　　处：《热带作物学报》2018年第2期300-307,共8页Chinese Journal of Tropical Crops

基　　金：海南省省属科研院所技术开发研究专项(No.KYYS-2016-10);农业部物种品种资源保护项目(No.151721301354051706);现代农业产业技术体系(No.CARS-33-26);海南自然科学基金面上项目(No.317191)

摘　　要：9-顺-环氧类胡萝卜素双加氧酶(9-cis-epoxycarotenoids dioxygenase,NCED)是ABA生物合成途径的关键限速酶,本研究利用RT-PCR技术从A4无核荔枝中分离得到NCED的3条c DNA序列(命名为LcNCED1、LcNCED2和LcNCED3)。通过同源比对和系统进化树分析,LcNCED基因编码的氨基酸序列与拟南芥等30个物种中的NCED氨基酸序列同源性都在65%以上,从而确定LcNCED属于NCED家族;通过与7条拟南芥及5条苹果NCED家族基因进行系统进化树分析,LcNCED2与调控苹果花瓣脱落MdNCED2、MdNCED4基因具有较高的同源性。Real-Time PCR表达分析发现,3个LcNCED基因在不同样品间表达存在显著差异,其中LcNCED2在各样品间的表达水平显著高于LcNCED1和LcNCED3,且LcNCED2表现出显著的组织特异性表达特点,花后15~75 d,LcNCED2在离区(果柄离层部位上下0.2 cm)中的表达量均高于种脐和果皮中,在花后45 d达到峰值,花后75 d稍有下降,基因表达的峰值略早于A4无核荔枝落果高峰期。而LcNCED1和LcNCED3在各组织样品中基因表达变化趋势规律不明显。上述结果表明,LcNCED2在离区中表达量变化比较符合A4无核荔枝生理落果趋势,推断LcNCED2基因可能与无核荔枝采前落果关系密切。9-cis-epoxycarotenoids dioxygenase （NCED） is the key enzyme for ABA synthesis in indirect pathway. Three full-length eDNA （temporarily named as LcNCED1, LcNCED2 and LcNCED3） encoding 9-cis-epoxycarotenoids dioxygenase were obtained from the litchi genome sequence resource （unpublished）, and were isolated with the techniques of RT-PCR. Phylogenetic tree analysis showed that amino acids sequence of LcNCED genes shared more than 65% identity with the sequence of NCED from other 30 plants; Compared with 7 NCED family gene in Arabidopsis thaliana and 5 NCED family gene in Malus domestiea, which showed high homology with A tNCED genes, LcNCED2 had the highest amino acids sequence identity with MdNCED2 and MdNCED4 which can control petals abscission. Real-time PCR results showed that LcNCED2 expression level was significantly higher than that of LcNCED1 and LcNCED3 in each sample. LcNCED2 showed significant characteristics of tissue specific expression, the expression in abscission zone presented higher than that in hilum and periearp. The expression level of LcNCED2 increased gradually during fruit development, and a rapid increase occurred in 45 DAF stages which is a little earlier than the fruit drop peak stage, The trend of expression of LcNCED2 in abscission zone was consistent with the trend of seedless litchi physiological fruit drop, the trend of expression of LcNCED1 and LcNCED3 were irregular. Thus, we considered that LcNCED2 gene might be the key gene invovled in fruitlet abscission in litchi.

关 键 词：荔枝 9-顺-环氧类胡萝卜素双加氧酶 基因克隆 基因表达 采前落果 

分 类 号：S435.33[农业科学—农业昆虫与害虫防治]