漾濞大泡核桃病程相关蛋白10基因JsPR10-1的克隆与表达分析

摘要

The activation and accumulation of pathogenesis-related protein (PR) 10 play predominant roles in the defense response against pathogens.Based on a Juglans sigillata expressed sequence tag (EST) encoding PR10,gene-specific primers were designed and used to amplify the full-length cDNA of a novel PR10 gene by rapid amplification of cDNA ends (RACE).This gene was named as JsPR10-1.JsPR10-1 was 776 bp in length with an open reading frame (ORF) of 483 bp,a 5'-untranslated region (UTR) of 74 bp,and a 3'-UTR of 219 bp,and the ORF encoded a protein with 160 amino acid residues.There was an intron of 124 bp in the genomic sequence of JsPR10-1.Homology analysis indicated that JsPR10-1 was homologous with PR10s from Quercus suber,Fagus sylvatica and Castanea sativa;moreover,JsPR10-1 clustered together with the PR10s from dicots in the phylogenetic tree.qRT-PCR analysis showed that the expression levels of JsPR10-1 were induced by four different plant signaling molecules,including salicylic acid,jasmonic acid,ethylene,and H2O2.In addition,after inoculation with Colletotrichum gloeosporioides,JsPR10-1 was sharply up-regulated with the highest expression level at 8 h.The JsPR10-1 gene was involved in the defense response against C.gloeosporioides.This experiment lays a theoretical foundation for revealing the mechanism of resistance in J.sigillata.%病程相关蛋白(pathogenesis-related protein,PR) 10的激活与积累在植物抗逆境胁迫中有非常重要的作用.根据漾濞大泡核桃(Juglans sigillata)编码PR10的EST (expressed sequence tag)序列设计引物,利用快速扩增cDNA末端技术,克隆得到PR10基因的全长cDNA序列,并命名为JsPR10-1.JsPR10-1全长cDNA为776 bp,含有483 bp的开放阅读框、74 bp 5'-非编码区以及219 bp 3'-非编码区,编码含有160个氨基酸的蛋白质.全长基因序列中含有1个124 bp的内含子.JsPR10-1编码的蛋白质与栎树(Quercus suber)、欧洲山毛榉(Fagus sylvatica)以及欧洲板栗(Castanea sativa)的PR10相似性较高,并且在PR10的系统进化树中与双子叶植物聚为一支.qRT-PCR分析结果表明,植物信号分子水杨酸、茉莉酸、乙烯以及过氧化氢处理均可诱导JsPR 10-1表达.在接种胶孢炭疽菌(Colletotrichum gloeosporioides)后,JsPR10-1的表达量迅速上升并在接种8h时达到最大值,表明JsPR10-1参与漾濞大泡核桃对胶孢炭疽菌的防卫反应.本研究为揭示漾濞大泡核桃抗性机制奠定了理论依据.