水稻白叶枯病菌hisF基因的克隆及功能初析  

Cloning and fuctional study of hisF gene from Xanthomonas oryzae pv.oryzae

作　　者：陈磊[1] 韩志成[1] 王晨[1] 杨万风[2] 钱国良[1] 胡白石[1] 刘凤权[1] 

机构地区：[1]南京农业大学农业部病虫监测与治理重点开放实验室,江苏南京210095 [2]连云港出入境检验检疫局,江苏连云港222042

出　　处：《南京农业大学学报》2009年第4期72-78,共7页Journal of Nanjing Agricultural University

基　　金：农业部公益性农业行业科技项目(NYHYZX07-056);国家863计划项目(2002AA245041)

摘　　要：根据黄单胞菌hisF基因的同源性设计简并引物,采用PCR方法从水稻白叶枯病菌(Xanthomonas oryzaepv.oryzae,Xoo)中克隆了hisF同源基因,命名为hisFXoo。NCBI网站蛋白质保守结构域搜索表明,HisFXoo属于组氨酸生物合成蛋白家族的一员,具有与其他HisF蛋白相似的结构。序列比较显示,该基因在黄单胞菌中是相对保守的。同源性搜索比较显示,与水稻条斑病菌(X.oryzaepv.oryzicola,Xooc)的hisF(登录号:DQ372107)同源性高达98%。通过同源重组的方法,构建了hisFXoo的插入突变体,在NA+Amp平板上筛选后,并且经过PCR及Southern杂交验证插入正确。在MMX基本培养基中生长测定发现,hisF突变体生长能力明显下降,证明hisF基因与Xoo生长代谢相关。但是,该突变体仍具有致病性,并且毒力与野生型菌株没有明显差异。A hisF homologue, designated hisFXoo, was cloned from the genomic DNA of Xanthomonas oryzae pv. oryzae （Xoo） with degenerated primers by PCR. NCBI blast search indicated that HisFXoo had a similar structure to that of other HisF proteins, and had conserved domain of histidine biosynthesis protein. Nucleotide sequence comparison showed that hisFXoo was conserved in the Xanthomonas genus. Homology search revealed that the hisFXooc was 98% similarities to hisF （Accession number： DQ372107） of X. oryzae pv. oryzicola （Xooc）. A hisFXoo disruption mutant was successfully generated by a single cross-over event, selected by Amp resistance on NA medium, and confirmed by PCR and Southern blot. The growth of the mutants was reduced compared to wild type strain when grown in MMX medium. The results suggest that hisFXoo is involved metabolism of Xoo. However, the mutant still had strong pathogenicity, and its virulence was not obviously different from that of wild type strain. Nevertheless, how hisF to regulate metabolism of Xoo, transcription and expression of genes involved in metabolism still need to be further studied.

关 键 词：水稻白叶枯病菌 hisF基因 突变体 生长代谢 

分 类 号：Q754[生物学—分子生物学] S435.111.1[农业科学—农业昆虫与害虫防治]