【作者】 汪川； 张朝武； 裴晓方； 刘衡川； 余倩； 许欣；

【Author】 WANG Chuan,ZHANG Chao wu,PEI Xiaofang,LIU Hengchuan,et al. Department of Medical Technology,West China School of Public Health,Sichuan University,Chengdu 610041,China

【机构】 四川大学华西公共卫生学院医检教研室；

【摘要】 目的对一株分离自酸奶经表型特征鉴定为德氏乳杆菌保加利亚亚种的菌株（wch9901）进行16SrDNA鉴定和系统发育分析。方法提取wch9901基因组DNA作为模板,以16SrDNA两端的保守序列作为PCR引物,扩增菌株的16S rDNA序列。在T₄连接酶的作用下,将扩增得到的16S rDNA序列插入克隆载体pGEM-T,构建重组质粒pGEE-wch9901 16S rDNA。重组质粒酶切鉴定合格后,交由测序公司对插入的16SrDNA序列进行测序。测序结果于GenBank中Blastn,并用软件Mega3.1构建系统发育树。结果 wch9901 16SrDNA序列与Lactobacillus delbrueckii subsp.bulgaricus中所有菌株的16S rDNA序列同源性均达96%以上;在系统发育树上wch9901单独形成一个分枝,位于Lactobacillus delbrueckii subsp.bulgaricus LGM2遗传分枝和Lactobacillus delbrueckii subsp.bulgaricus DL2所在的独立小遗传簇及Lactobacillus delbrueckii subsp.bulgaricus JSQ所在的独立小遗传簇构成的分枝之间,并与Lactobacillus delbrueckii subsp.bulgaricus LGM2遗传分枝距离最近。结论 wch9901属于Lactobacillus delbrueckii subsp.bulgaricus;wch9901与Lactobacillus delbrueckii subsp.bulgaricusLGM2具有最近的亲缘关系。更多还原

【Abstract】 Objective For being further applied and studied,one strain of Lactobacillus delbmeckii subsp.bulgaricus （wch9901） separated from yoghourt which had been identified by phenotype characteristic analysis was identified by 16S rDNA and phylogenetic analyzed.Methods The 16S rDNA of wch9901 was amplified with the genomic DNA of wch9901 as template,and the conservative sequences of the 16S rDNA as primers.Inserted 16S rDNA amplified into clonal vector p GEM- T under the function of T₄ DNA ligase to construct recombined plasmid p GEM-wch9901 16 S rDNA.The recombined plasmid was identified by restriction enzyme digestion,and the eligible plasmid was presented to sequencing company for DNA sequencing.Nucleic acid sequence was blastn in GenBank and phylogenetic tree was constructed using neighbor joining method of distance methods by Mega3.1 soft.Results Results of blastn showed that the homology of 16S rDNA of wch9901 with the 16S rDNA of Lactobacillus delbrueckii subsp.bulgaricus strains was higher than 96%.On the phylogenetic tree,wch9901 formed a separate branch and located between Lactobacillus delbrueckii subsp.bulgaricus LGVE ewlution branch and another ewlution branch which was composed of Lactobacillus delbmeckii subsp.bulgaricus EX2 ewlution cluster and Lactobacillus delbmeckii subsp.bulgaricus JSQ ewlution cluster.The distance between wch9901 ewlution branch and Lactobacillus delbmeckii subsp.bulgaricus LGVE ewlution branch was the closest.Conclusion wch9901 belonged to Lactobacillus delbmeckii subsp.bulgaricus.wch9901 showed the closest ewlution relationship to Lactobacillus delbmeckii subsp.bulgaricus L GM2.更多还原