一株羊源单核细胞增生性李斯特菌的分离与鉴定一马光强.doc

一株单增李斯特菌的分离 马光强1(，1，文 明12，王开功12，程振涛12*，周碧君12* 贵州大学动物科学学院，贵州 贵阳 550025550025） 摘要:本研究旨在了解一株单核细胞增生李斯特菌贵州分离株的理化特性及遗传变异情况从贵州省某分离李斯特菌，对该菌进行鉴定。Lm的hly基因引物，以分离菌株基因组DNA为模板，扩增hly基因，经回收纯化与MD19-T载体连接后转化感受态细胞DH5α、双酶切鉴定测序及序列。结果分离培养特性、菌落形态、菌体形状特征、生理生化特征均与文献报道李斯特菌。从该分离菌株中扩增到850bp的Lm的特异性DNA片段，其序列与GenBank地方株核苷酸同源性在.1%-99.7%之间，其中与的加拿大瑞士参考株同源性最高，均达到99.7%，与其他参考菌株同源性李斯特菌hly基因。 关键词:单增李斯特菌；分离鉴定；hly基因；Isolation and Identification of a Listeria monocytogenes in Caprine MA Guang-qiang1, LIU Li-juan1,2,WEN Ming1,2,WANG Kai-gong1,2,CHENG Zhen-tao1,2*, ZHOU Bi-jun1,2* （1. College of Animal Science, Guizhou Uiversity, Guiyang 550025, China; 2. Key laboratory of animal disease and veterinary public health in guizhou province, Guiyang 550025, China） Abstract: This study aims to understand the characterization and genetic variation of a Listeria monocytogenes (Lm) in Guizhou, applying the laboratory isolation and culture techniques isolated a Lm from the tissue samples of a infected case in a Caprine farm in Pan County of Guizhou，and use the traditional test methods for its identification. A pair of specific primer designed and synthesized to amplify the sequence of hly gene in isolated Lm strain’s genomic DNA. The hly was cloned into pMD19-T vector, positive colonies were sequenced and digested by restriction endonuclease after transforming, Sequence analysis for it then. The results showed that a suspected Lm have been isolated from a suspected Caprine case, and its cultural characteristics, colony morphology, shape features, and Physiological-biochemical characteristics are the same as the Lm in literature reports. A specific DNA fragment about 850bp of Lm was amplified, and the nucleotide homology between 39.1% -99.7% compared with part of local strains in GenBank, Canada and Switzerland（99.7% ）isolated in onset of animals is the highest, very low with the other strain isolated from other ways, which is further confirms from the molecular level that a Lm has been isolated, and Large ge