猪“高热病”源嗜麦芽窄食单胞菌16S rDNA基因的克隆和系统发育分析

Cloning and Phylogenetic Analysis of 16S rDNA Gene of Stenotrophomnas maltophilia Derived from Swine High Fever Cases

根据NCBI中已报道的嗜麦芽窄食单胞菌16S rDNA基因序列设计引物,对两株来源于猪"高热病"病料中的疑似嗜麦芽窄食单胞菌PSM-5、PSM-6进行PCR扩增。将扩增产物克隆至pMD18-T载体上,并转化到DH5α中。抽取质粒模板,进行PCR和双酶切鉴定并测序,从分子水平上予以鉴定。两株菌的16S rDNA基因扩增片段的核苷酸序列(GenBank登录号为GQ267816和GQ267817)与已经报道的嗜麦芽窄食单胞菌分离株的16S rDNA序列的相似性均在96.78%以上,最高达99.93%。本试验对通过16S rDNA鉴定嗜麦芽窄食单胞菌提供一定参考,同时对猪"高热病"的诊断治疗奠定细菌学方面的基础,对其发病机制的探讨提供依据。

According to the published Stenotrophomnas maltophilia 16S rDNA gene sequences in NCBI,we designed a pair of primers.Two suspected Stenotrophomnas maltophilia strains（PSM-5,PSM-6） were amplified by PCR using the primer.The PCR products were cloned into pMD18-T vectors and transformed into competent cell DH5α.The recombinant plasmid was identified by PCR and restriction enzyme and then sequenced,identifying from the molecular level.The nucleotide identity of 16S rDNA gene（GQ267816,GQ267817） amplified from PSM-5 or PSM-6 strain was of above 96.79%,up to 99.93% compared to those of the Stenotrophomonas maltophilia,found in the GenBank,which were isolated from different environment.The experiment provides a reference to identify the Stenotrophomonas maltophilia via the 16S rDNA gene,while it lays the foundation of bacteriology for the diagnosis and treatment of the swine high fever,and it makes further discussion of the pathogenesis as well.