西瓜噬酸菌pilN基因功能分析  

Analysis of the pilN gene in Acidovorax citrulli

作　　者：王健超[1] 王敏鑫[1] 代园凤[1] 袁云云[1] 田艳丽[1] 武珍[1] 陈斌[1] 胡白石[1] 

机构地区：[1]南京农业大学植物保护学院/农作物生物灾害综合治理教育部重点实验室,江苏南京210095

出　　处：《南京农业大学学报》2013年第2期51-57,共7页Journal of Nanjing Agricultural University

基　　金：国家重点基础研究发展计划项目(2009CB119200);全国科技支疆计划项目(200840002-08);国家公益性行业(农业)科研专项(201003066)

摘　　要：为了分析菌毛组装蛋白家族基因对瓜类细菌性果斑病菌致病性的影响,以xjl12菌株为背景构建转座子(Tn5)插入的突变体文库,通过浸种处理和子叶注射接种方法筛选致病性相关突变体,通过两亲交配获得突变株,并对所得的突变体、野生型及互补等多个菌株进行致病性、过敏性反应、游动性等相关表型进行测定,用反转录PCR(qRT-PCR)方法验证xjl12和突变体中hrpG、hrcV、celA表达量的差异。结果表明:突变体文库中筛选得到1株致病力明显下降的突变体Δxj-7,经亚克隆鉴定为pilN基因,其功能主要与菌毛(pili)生物合成相关。两亲交配后所得突变菌株ΔpilNac相较于野生型,其致病性、游动性、胞外纤维素酶活性降低,同时丧失烟草过敏性反应。qRT-PCR试验结果显示,Ⅲ型分泌系统的主要调控基因以及纤维素酶生物合成基因在ΔpilNac中的转录水平明显下调。证明菌毛生物合成相关基因pilN对致病性有重要的调控作用。To investigate functions of fimbrial assembly family gene in Acidovorax citrulli （Ac）, we generated a transposon （Tn5） mutant library on the background of strain xjll2 and screened it for reduced virulence by seed-transmission assays and injection inoc- ulation with melons. The gene insertional mutant ApilNac was generated by biparental mating from the wild-type strain xjll2, meanwhile cell morphology, motility and pathogenicity in host and hypersensitive response （HR）in nonhost tobacco were determined. Here,we got one strain Axj,7 with reduced virulence. This Tn5 mutant was impaired in pilN, which was involved in fimbrial assembly family protein and played a part in the biosynthesis of type IV pili. The pathogenicity on seed leaf was obviously attenuated in ApilNac compared to wild type. There was significant difference in bacterial growth in MMX medium, production of cellulose and induction of HR between mutant（△pilNac） and wild type. In addition, these results of qRT-PCR demonstrated that the transcription level of hrpG, hrcV and celA were down-regulated in ApilNac. The complementation strain restored the phenotypes of piIN gene partially or completely. This study showed that the △pilNac mutation had influence not only in growth, but also in pathogenicity in Ac.

关 键 词：西瓜噬酸菌 转座子 菌毛组装蛋白 致病性 

分 类 号：S436.42[农业科学—农业昆虫与害虫防治]