摘要
利用含有无启动子的Km抗性基因的转座子mariner对胡萝卜软腐果胶杆菌胡萝卜亚种(Pectobacterterium carotovorum subsp.carotovorum,Pcc)菌株PccS1进行转座诱变,建立突变体文库,以白花马蹄莲组织提取液为诱导物,诱导突变菌株,鉴定突变、并验证。筛选获得1株受白花马蹄莲组织提取液诱导的突变株PM5,经亚克隆鉴定,转座子插入位点为辅酶Q氧甲基转移合成酶ubiG基因(Ubiquinone biosynthesis O-methyltransferase)。利用同源重组获得该基因的缺失突变体△ubiG。与野生型相比,△ubiG对不同寄主的致病性均减弱(黄花马蹄莲和白花马蹄莲上的病斑面积减半,大白菜的病斑面积仅为1/5);突变菌株菌体沉降速率明显加快,生物膜产量降低约50%,无胞外蛋白酶产生。此外,△ubiG对链霉素和新霉素抗性分别比野生型增强了2.0倍和26.5倍,突变菌株的相关功能经互补可以部分恢复。以上结果说明,ubiG基因可被白花马蹄莲诱导并参与胡萝卜软腐杆菌的致病作用。
A mutant library was constructed by rnar/ner, a promoter-free transposon, in Pectobacterterium carotovorum subsp, carotovorum （Pce） strain PecS1. Only one mutant PM5 was obtained after induction and identification. The transposon insertion site was identified as an ubiG gene coding ubiquinone biosynthesis O-methyhransferase. The gene deletion mutant Au- big was further constructed by homologue recombination. Compared with the wild type PccS1, the pathogenicities of AubiG to various host plants were weakened, with halved lesion areas in colored calla and caUalily, and 20% lesion area in Chinese cabbage. AubiG presented accelerated sedimentation, halved biofilm and zero extracellular protease production. The resistances of AubiG to streptomycin and neomycin were in- creased by 2. 0 and 26.5 times compared to the wild type, re- spectively. The pathogenicities of the mutant were restored partly in complemented strain. In conclusion, the ubiG gene was induced by Z. aethiopica and played an important role in pathogenesis of Pcc.
出处
《江苏农业学报》
CSCD
北大核心
2013年第6期1304-1312,共9页
Jiangsu Journal of Agricultural Sciences
基金
江苏省农业科技支撑计划项目(BE2013385)
江苏省农业科技自主创新基金项目[CX(12)3016]
关键词
胡萝卜软腐果胶杆菌
mariner转座子
诱导表达
基因敲除
功能分析
Pectobacterterium carotovorum subsp. Carotovorum
mariner transposons
inducible expression
geneknockout
functional analysis
