【摘要】 根据青枯菌(Ralstonia solanacearum)与群体猝灭相关aac基因序列设计PCR引物,扩增并克隆了aac基因,将庆大霉素基因(Gmr)插入aac基因内部,亚克隆至自杀质粒pDS132中,构建了aac基因重组自杀质粒pDS-aac-Gm。将自杀质粒电转化至青枯菌GMI1000菌株中,通过体内同源重组,置换其野生型的aac基因。通过三步筛选法和PCR扩增鉴定,筛选获得了具有庆大霉素抗性的青枯菌aac基因突变株(GMI1000-m)。土壤接种番茄青枯菌结果显示,突变株GMI1000-m的致病性较野生型GMI1000明显下降。证明aac基因在青枯菌致病过程中具有重要作用。更多还原

【Abstract】 According to the sequence of aac gene associated with quorum-quenching of Ralstonia solanacearum, the PCR primers were designed to amplify and clone aac gene. The aac gene was mutated by inserting gentamicin-3-acetyltransferase gene (Gmr gene) and the mutated gene was subcloned into suicide vector pDS132, named pDS-aac’-Gm. Then the vector pDS-aac-Gm was introduced into R. solanacearum GMI1000 strain and aac gene mutant was generated by homologous recombination. The aac gene mutated strain with Gmr (GMI1000-m) was selected by three-step methods and aac inserted by Gm gene on the genome was determined by PCR. The result of tomato inoculation test showed that the GMI 1000-m strain were much less virulent on tomato than the wild-type GMI1000, which indicated that the aac gene is a very important factor in the pathogenesis of R. solanacearum.更多还原