野核桃低温响应转录因子JcICE1基因的克隆与分析  

Gene cloning and analysis of inducer of CBF expression 1 gene(ICE1)fromJuglanscathayensis

作　　者：陈尘 张燕[1] 高书宝[1] 岳明 CHEN Chen;ZHANG Yan;GAO Shubao;YUE Ming(Shaanxi Engineering Research Centre for Conservation and Utilization of Botanical Resources,Xi'an Botanical Garden of Shaanxi Province(Institute of Botany of Shaanxi Province),Xi'an 710061,China)

机构地区：[1]陕西省植物资源保护与利用工程技术研究中心,陕西省西安植物园(陕西省植物研究所),西安710061

出　　处：《植物生理学报》2024年第11期1697-1706,共10页Plant Physiology Journal

基　　金：陕西省科学院科技计划项目(2019k-07)。

摘　　要：低温胁迫严重影响了野核桃(Juglans cathayensis)生长发育。为分析低温响应转录因子(inducer of CBF expression 1)ICE1基因在野核桃应答低温胁迫中的关键作用,本研究基于转录组数据,以野核桃叶片为材料,设计特异性引物克隆了野核桃ICE1基因;采用生物信息学技术对其编码蛋白的基本性质、系统进化进行分析;并利用实时荧光定量PCR(qRT-PCR)检测了JcICE1基因应答低温胁迫和茉莉酸甲酯处理时的表达模式。结果表明:(1)JcICE1基因包含一个1590 bp的开放读码框,编码530个氨基酸,预测蛋白分子质量57.58 kDa,等电点为5.35,为不稳定亲水蛋白;(2)JcICE1蛋白含有典型的b HLH特征结构域,系统进化分析结果表明JcICE1与核桃的ICE1亲缘关系较近;(3)JcICE1启动子区域含有应答低温、响应环境胁迫的顺式所用元件;qRT-PCR分析发现,低温和茉莉酸甲酯处理均能显著诱导JcICE1的表达:低温处理36 h后JcICE1表达量是对照的5.12倍(P<0.01);茉莉酸甲酯处理8 h时JcICE1的表达量最高,为对照的2.44倍(P<0.01)。本研究为野核桃应答低温胁迫的分子机制和通过基因工程培育耐寒核桃品种提供支撑。Low temperature stress seriously affects the growth and development of Juglans cathayensis.To investigate the key function of the low temperature-responsive transcription factor,inducer of CBF expression 1 gene(ICE1)in J.cathayensis,this study cloned the JclCE1 gene through searching transcriptome data.Then the fundamental characteristics and phylogenetic evolution of the encoded protein were examined using bioinformatics tools.Additionally,the expression pattern of JclCE1 gene in reaction to low temperature and MeJA treatment was assessed by real-time fluorescence quantitative PCR(qRT-PCR).The results indicated that JciCE1 gene comprises an open reading frame of 1590 bp,encoding a protein of 530 amino acids.This protein has a predicted molecular mass of 57.58 kDa and an isoelectric point of 5.35,characterizing it as an unstable hydrophilic protein.Furthermore,JcicE1 protein contains typical bHLH domains.The phylogenetic analysis revealed a close relationship between JclICE1 and JrlCE1 in walnut.The promoter region of JclCE1 contained cis-elements responsive to low temperature and environmental stress.qRT-PCR analysis showed that both low temperature and MeJA treatments significantly upregulated JclCE1 expression,with a 5.12-fold and 2.24-fold increase compared that of the control after 36 hours of low temperature treatment and 8 hours of MeJA treatment,respectively(P<0.01).This study offers evidence for the molecular mechanism in the response of J.cathayensis to low temperature stress,as well as the potential for developing cold-resistant walnut cultivars using genetic modifications techniques.

关 键 词：低温响应转录因子 基因克隆 亚细胞定位 表达模式 野核桃(Juglans cathayensis) 

分 类 号：S664.1[农业科学—果树学]