富民枳WRKY基因家族的鉴定和干旱胁迫表达分析  

Identification of WRKY gene family and analysis of drought stress expression in Poncirus polyandra

作　　者：张珊珊 阮孙美 杨文忠 ZHANG Shanshan;RUAN Sunmei;YANGWenzhong(Yunnan Academy of Forestry and Grassland Sciences/Yunnan Key Laboratory of Conservation and Breeding of Rare and Rare Forest Plants/National Forestry and Grassland Administration,Kunming 650201,Yunnan,China)

机构地区：[1]云南省林业和草原科学院·国家林业和草原局云南珍稀濒特森林植物保护和繁育重点实验室,昆明650201

出　　处：《果树学报》2025年第3期508-525,共18页Journal of Fruit Science

基　　金：国家自然科学基金地区基金项目(31960278);云南省林业和草原科学院基金项目(MS2019-08,MS2019-09)。

摘　　要：【目的】富民枳(Poncirus polyandra)为柑橘属近缘种,是嫁接时广泛使用的抗性砧木,属云南省特有的极小种群野生植物。WRKY家族是植物中特有且最大的转录因子之一,在植物应对非生物胁迫过程中发挥着重要的作用。研究富民枳WRKY基因家族,为解析富民枳WRKY基因功能以及筛选抗旱基因提供理论依据。【方法】基于富民枳基因组数据,利用生物信息学方法对富民枳WRKY基因家族成员进行全基因组鉴定,系统分析该家族成员的理化性质、序列特征、系统进化关系、染色体定位、共线性、基因结构和顺式作用元件等以及qRT-PCR分析干旱胁迫下的表达模式。【结果】在富民枳基因组中共鉴定到46个WRKY家族基因,命名为PpWRKY1~PpWRKY46,染色体定位分析显示,WRKY家族基因不均匀地分布在9条染色体上。亚细胞定位预测表明,富民枳WRKY家族成员主要定位于细胞核上。WRKY结构域多序列比对分析显示,只有Ⅱc亚组的PpWRKY15和PpWRKY21的典型七肽序列发生单氨基酸变异。共线性分析结果表明,富民枳和枳有88对共线性基因;片段复制是PpWRKY基因家族扩张的主要驱动力。顺式作用元件分析表明,PpWRKY家族基因启动子区域含有大量胁迫类和激素类响应顺式作用元件。基于富民枳近缘种枳的WRKY家族基因在干旱胁迫下的转录组数据,在富民枳中筛选到PpWRKY8、PpWRKY12、PpWRKY13、PpWRKY19、PpWRKY23、PpWRKY27、PpWRKY28、PpWRKY30和PpWRKY41共9个同源抗旱候选基因,最后通过qRTPCR分析表明,这9个基因均受干旱胁迫诱导显著表达。蛋白互作预测结果表明,有12个PpWRKY蛋白可能通过MYB转录因子介导ABA信号参与调控气孔运动来影响植物的耐旱性。【结论】鉴定到9个可能参与调控干旱胁迫的PpWRKY基因,为解析WRKY基因在富民枳干旱胁迫响应中的功能提供新信息,并为培育耐旱柑橘品种提供抗旱基因资源。【Objective】Poncirus polyandra,a relative species of citrus,is a type of resistant rootstock widely used in grafting.It is also a plant species with extremely small populations in Yunnan.The WRKY family,as one of the largest families of transcription factors in plants,plays a vital role in plant response to abiotic stresses such as drought,cold and salt.However,the current understandings of WRKY genes in P.polyandra are extremely limited.The purpose of this study is to analyse the function of WRKY genes and screen out the genes with potential drought resistances in P.polyandra.【Methods】Based on the genome-wide data of P.polyandra,WRKY genes were identified by HMMER,NCBICDD and SMART searches.Comprehensive analyses were systematically performed using bioinformatics methods,including the physicochemical properties,sequence characteristics,phylogenetic relationships,chromosome localization,collinearity,gene structure and cis-acting elements of the PpWRKY family members.In order to verify the drought resistance function of WRKY gene family members,oneyear-old P.polyandra seedlings were used as experimental materials.In the drought stress experiments,the seedlings were treated with 20%PEG-6000 solution.The leaves were respectively collected after 0 h,3 h,6 h,12 h and 24 h,and stored in the refrigerator at-80℃.Real-time fluorescence quantitative analysis was carried out to study the effects of drought stress on the expression patterns of these genes.【Results】A total of 46 WRKY family genes were identified in the genome of P.polyandra and named PpWRKY1-PpWRKY46.The results showed that the length of PpWRKY protein ranged from 116 to 1103 amino acids.Among them,the molecular weight of PpWRKY21 was the lowest(13213.48 kDa),whereas PpWRKY23 had the highest molecular weight(120198.96 kDa).The protein isoelectric points(pI)ranged from 4.93(PpWRKY24)to 9.8(PpWRKY17),and 61%of them was lower than 7.0,indicating that most of the proteins were acidic.Subcellular localization results exhibited that PpWRKY23 was locate

关 键 词：富民枳 WRKY基因 干旱胁迫 生物信息学 表达分析 

分 类 号：S666[农业科学—果树学]