华丽龙胆二氢黄酮醇-4-还原酶GsDFR基因的克隆及表达模式分析  

Cloning and Expression Pattern Analysis of Dihydroflavonol 4-Reductase Genes GsDFR in Gentiana sino-ornata

作　　者：方颖 赵海平 孔祥莹 衡蒙 金雪花 Fang Ying;Zhao Haiping;Kong Xiangying;Heng Meng;Jin Xuehua(Faculty of Architecture and City Planning,Kunming University of Science and Technology,Kunming,650500)

机构地区：[1]昆明理工大学建筑与城市规划学院,昆明650500

出　　处：《分子植物育种》2023年第6期1820-1827,共8页Molecular Plant Breeding

基　　金：国家自然科学基金项目(31760587,31560568)资助。

摘　　要：二氢黄酮醇-4-还原酶(DFR)是花青素苷合成过程中的关键酶。为研究华丽龙胆(Gentiana sino-ornata) DFR与花色形成的关系,将其花冠不同开放阶段分为H1~H5阶段,采用RT-PCR技术首次从华丽龙胆中克隆得到GsDFR全长序列,对其进行生物信息学和表达模式的分析。结果显示GsDFR包含1 125 bp的开放阅读框(OFR),编码375个氨基酸。结构分析显示,GsDFR属于NADB_Rossmann超家族,具有典型的DFR蛋白(PLNO02650)功能结构域,含有“TGASGYI”和“TVDVQEQQKPVYDENDWSDLDFINSH”两个典型的结构域,以及FR_SDR_e的特征位点。系统进化树分析表明Gs DFR与橙花龙胆(Gentiana lutea var.aurantiaca)亲缘性最近。采用实时荧光定量PCR分析GsDFR在根、茎、叶和花冠中的表达,发现GsDFR在根、茎、叶和花冠中均有表达,其中花冠的表达量最高,根茎中微量表达。在花冠H2阶段GsDFR表达量达到最高,随后降低。结果表明,华丽龙胆蓝色花冠中的GsDFR高表达可能会导致花青素苷的积累,使华丽龙胆花冠呈更艳丽的蓝色。本研究结果为揭示GsDFR在华丽龙胆蓝色花冠中花青素苷的积累的作用提供了理论基础。Dihydroflavanol-4 reductase(DFR)is a key enzyme in the synthesis of anthocyanins.In order to study the relationship between DFR and flower color formation,different corolla opening stages of Gentiana sino-ornata were divided into H1~H5 stages.A full-length sequence of GsDFR was cloned from Gentiana sino-ornata by RT-PCR for the first time,and the bioinformatics and expression pattern of the sequence were analyzed.It was found that GsDFR contained 1125 bp open reading frame(OFR),encoding 375 amino acids.Structure analysis showed that GsDFR belongs to NADB_Rossmann super family,with functions of typical DFR(PLNO02650)protein structure domain,containing"TGASGYI"and"TVDVQEQQKPVYDENDWSDLDFINSH"the structure of the two typical domain and the characteristic site of FR_SDR_e.Phylogenetic tree analysis showed that GsDFR was the most closely related to Gentiana lutea var.aurantiaca.RT-qPCR was used to analyze the expression of GsDFR in roots,stems,leaves and corolla.The results showed that GsDFR was expressed in roots,stems,leaves and corolla,with the highest expression in corolla and a trace expression in rhizome.GsDFR expression reached the highest level at H2 stage of corolla and then began to decrease.The results reflected that the high expression of GsDFR in the blue corolla of Gentiana sino-ornata might lead to the accumulation of anthocyanins,which would make the corolla of Gentiana sino-ornata more colorful blue.These results provide a theoretical basis for revealing the accumulation of GsDFR anthocyanins in the blue corolla of Gentiana sino-ornata.

关 键 词：华丽龙胆 蓝色花 GsDFR 基因克隆 表达分析 

分 类 号：Q943.2[生物学—植物学]