早酥梨抗黑星病相关基因PbzsREMORIN的克隆及功能分析  

Cloning and Functional Analysis of PbzsREMORIN Gene Related to Venturia nashicola Resistance in‘Zaosuli’Pear

作　　者：付镇芳[1,2] 姚春潮 张朝红 王跃进 

机构地区：[1]西北农林科技大学园艺学院农业部西北地区园艺作物生物学与种质创制重点实验室 [2]西北农林科技大学旱区作物逆境生物学国家重点实验室,陕西杨凌712100

出　　处：《园艺学报》2012年第1期13-22,共10页Acta Horticulturae Sinica

基　　金：国家‘863’计划项目(2007AA10Z182);陕西省重点科技专项(2006kz05-G4)

摘　　要：以高抗黑星病的早酥梨为试材,利用从梨抗黑星病抑制消减文库中筛选出病原菌诱导特异表达基因片段,通过RACE技术克隆获得其全序列,命名为PbzsREMORIN(GenBank登录号为HQ901373)。该基因全长897bp,开放阅读框(ORF)597bp,编码198个氨基酸;生物信息学分析表明该基因具有remorin家族保守结构域;实时荧光定量RT-PCR结果表明,该基因受梨黑星病病原菌的诱导;用基因枪法将基因与GFP的融合蛋白转化到洋葱鳞茎表皮细胞中,瞬时表达显示remorin蛋白定位于细胞膜、细胞质和细胞核中;将目的基因转入烟草品种NC89,共获得27个转基因烟草株系;离体叶片接种烟草青枯病病原菌结果表明,过量表达该基因增强了NC89对烟草青枯病病原菌的抗性。该基因可能在植物与病原菌互作过程中起重要作用。Abstract A pathogeninducible expression gene fragment, from a previously constructed Suppression Subtractive Hybridization （SSH） cDNA library of ＇Zaosuli＇ pear, was selected to obtain the full length of cDNA by RACE technique. The obtained cDNA, designated PbzsREMORIN, was 897 bp in length with a 597 bp open reading frame （ORF） and encoded 198 amino acid residues. Bioinformatics analysis indicated that PbzsREMORIN possesses a conserved domain of remorin family genes. The result from realtime RTPCR assay revealed that the target gene was induced by Venturia nashicoIa. Moreover, the PbzsREMORIN gene was fused with GFP and transferred into onion epidermal cells via particle bombardment. Transient expression assay demonstrated that the PbzsREMORIN protein was located in thecell membrane, cytoplasm and nucleus. The PbzsREMORIN gene was transferred into Nicotiana tabacum ＇NC89＇ and totally obtained 27 transgenic lines. Analysis of invitro transgenic tobacco leaves with Ralstonia solanacearum suggested that overexpression of PbzsREMORIN gene in tobacco increased resistance to R. solanaeearum, which indicated that the gene may play a critical role in plantpathogen interaction.

关 键 词：梨 黑星病 remorin 基因表达 转基因烟草 

分 类 号：S661.2[农业科学—果树学]