椰酵假单胞菌与唐菖蒲伯克霍尔德菌16S～23S rRNA基因间区序列的比较研究

Study on Comparison of 16S～23S rRNA Gene ISR Sequence of Pseudomonas cocovenenans subsp farinofermentans Strains and Burkholderia gladioli Strains

目的研究比较我国分离的椰酵假单胞菌与唐菖蒲伯克霍尔德菌不同致病型菌株的16S～23S rRNA基因间区序列,分析不同菌株基因水平的差别,并阐述其系统发育关系。方法设计2对伯克霍尔德菌属特异引物,扩增16S～23SrRNA基因间区序列。应用分子生物学软件,对3株分别代表唐菖蒲伯克霍尔德菌3种不同致病型的国际参考菌株(ATCC10248、NCPPB947、NCPPB3580)、1株伯克霍尔德菌属B.phenazinium种代表株(LMG2247)及8株从我国不同地域、不同食物样品中分离的椰酵假单胞菌(HN2y、Co14、Co8、Co36、Sx8801、90-3、56(2)、56(2))的16S～23SrRNA基因序列进行测定,并与核酸数据库(GenBank)中相关菌株序列进行比较分析。结果通过不同菌株16S～23SrRNA间区序列的分析比较,发现分离的椰酵假单胞菌米酵菌酸产毒株存在2个序列高可变区及特异基因序列,阐述了我国椰酵假单胞菌与唐菖蒲伯克霍尔德菌的系统发育关系,将DNA测序结果在核酸数据库中注册,并绘制了系统发育进化树。结论该研究结果为进一步鉴定椰酵假单胞菌米酵菌酸产毒株,从分子水平探讨其产毒机制及系统发育关系提供了新的、可靠的理论依据和技术支撑。

Objective To study and analyze the differences among the strains in the genus Burkholderia at gene levels by comparing 16S - 23S rRNA gene intergenic spacer region （ISR） sequences of Pseudomonas cocovenenans subsp farinofermentans strains isolated in Chinese foods and Burkholderia gladioli model strains. The phylogenetic relationships of them were expounded. Method Two pairs of specific primers for the genus Burkholderia were designed and 16S - 23S rRNA gene ISR sequences were amplified for the strains. 16 - 23S rRNA gene ISR sequences of 3 international reference strains representative of the three different pathogenic strains of Burkholderia gladioli （ATCC10248, NCPPB 947, NCPPB3580）, and 1 strain of B. phenazinium （LMG2247） and 8 strains of Pseudomonas cocovenenans subsp farinofermentans isolates （HN2y, Co14, Co8, Co36, Sx8801, 90-3, 56 （2）, 56 （2）） from the different samples of food-poisoning in China were determined. The total sequences were compared and analyzed with the relative strain sequences in the GenBank by the molecular biological analyzing software. Results Two highly variable ISR regions and the DNA specific sequences associated with isolate strains produced the Bongkrekic acid were found. The results of the DNA sequences were submitted to the GenBank. The phylogenetic tree of the isolates and the Burkholderi gladioli were illustrated. Conclusion New and reliable theory and technology supports were provided for the further study of the identification of Pseudomonas their mechanisms in molecular levels and subsp farinofermentans isolates produced Bongkrekic acid, research into their phylogenetic relationships