十字花科黑腐病菌应急反应相关基因spoT_(Xcc)对T3SS基因调控作用的分析

Analysis of regulatory relationship between stringent response related gene spoT_(Xcc)and T3SS genes in Xanthomonas campestris pv.campestris

通过反向遗传学方法和半定量RT-PCR方法,研究了十字花科黑腐病菌中被注释的应急反应相关基因spoT_(Xcc)与Ⅲ型分泌系统(T3SS)基因的调控关系。基于自杀质粒pK18mob同源整合方法构建了十字花科黑腐病菌spoT_(Xcc)基因(XC_0956)的非极性整合突变体,检测了突变体的多种表型,发现spoT_(Xcc)突变体延迟了在非寄主植物辣椒ECW-10R上引发的过敏反应(HR)。利用GUS(β-葡糖苷酸酶)活性检测、原位组织染色和半定量RT-PCR的方法,发现T3SS相关基因在spoT_(Xcc)基因的非极性整合突变体中表达量降低,表明十字花科黑腐病菌中被注释的应急反应相关基因spoT_(Xcc)显著正调控T3SS相关基因的表达。

The regulatory relationship between the stringent response related gene spoTxcc and type III secretion system （T3SS） related genes in the bacterial phytopathogen Xanthomonas campestris pv. campestris was investi- gated by using reversed genetics method and semi-quantitative RT-PCR. A non-polar mutant of spoTxcc （XC_ 0956） was generated by suicide plasmid pK18mob mediated homologous integration and various phenotypes of the mutant were studied. It was found that the spoTxcc mutant delayed the induction of hypersensitive response on the non-host plant pepper ECW-10R. The expression of T3SS genes were down-regulated in spoTXcc gene mutant detected by GUS activity tests, semi-quantitative RT-PCR and staining of tissue methods, indicating that the stringent response related gene spoTXcc positively regulated the T3SS related genes in X. campestris pv. campestris.