中国畜牧兽医2019，Vol.46Issue(1)：45-53,9.DOI:10.16431/j.cnki.1671-7236.2019.01.006
食源性单增李斯特菌inlA/inlB/inlC基因缺失株的构建及其生物学特性分析
Construction and Characterization of inlA/inlB/inlC Genes Deleted Mutant Strain of Food-borne Listeria monocytogenes
摘要
Abstract
In order to evaluate the effect of inlA/inlB/inlCgenes on biological characteristics of Lm, the inlCgene deletion mutant was constructed by fusion PCR method, and then pKSV7-△inlC shuttle vector was constructed, which was transformed into competent cells Lm681-△inlAB.Homologous recombination was conducted using temperature (42 ℃) and chloramphenicol (10μg/mL) resistance to achieve the inlCgene deletion strain, and then the homologous recombination was screened for identification and some biological characteristics were studied.PCR and sequencing results confirmed that Lm681-△inlABC was successfully obtained.The results showed that the growth characteristics of the deletion mutants were the same as the wild-type strain Lm681 and there was no significant difference.The hemolysis activity of the mutant strains were consistent with wild-type strain Lm681.In the mouse infection experiment, the mortality rate of wild-type strain Lm681, Lm681-△inlAB and Lm681-△inlABC were 80％ (8/10) , 60％ (6/10) and40％ (4/10) , respectively.Results of mice virulence determination showed that the median lethal dose of wild-type strain, Lm681-△inlAB and Lm681-△inlABC were 4.36×10`4, 1.35×106 and 2.95×107 CFU, respectively.Furthermore, the colonization ability of the inlA/inlB/inlCgenes deleted strain was extremely significantly lower than wild-type strain in liver, spleen and brain (P<0.01) .Collectively, these results indicated that inlA/inlB/inlCgenes might play an important role in pathogenicity of Lm infection and provided scientific basis for the study on the mechanism in host cell invasion by Lm.关键词
单增李斯特菌/内化素基因/基因突变株/生物学特性Key words
Listeria monocytogenes/internalin gene/gene mutant/biological characteristics分类
农业科技引用本文复制引用
任静静,杨铭伟,陈云飞,尹华,王欣雨,李蓓蓓,蒋建军,王鹏雁..食源性单增李斯特菌inlA/inlB/inlC基因缺失株的构建及其生物学特性分析[J].中国畜牧兽医,2019,46(1):45-53,9.基金项目
国家自然科学基金项目(31260606) （31260606）
