单核细胞增生性李斯特菌fbpa基因敲除菌株的构建

摘要

Objective To construct an fbpA-deletion mutant of Listeria monocytogenes. Methods The fbpA gene and its upstream, downstream genes of Listeria monocytogenes were cloned into plasmid pCR Ⅱ. The upstream and downstream fragments were ligated into the pAULA using restriction enzyme as pAULA-ΔfbpA. To achieve allelic exchange, pAULA-ΔfbpA was introduced into Listeria monocytogenes by electroporation. The mutant was confirmed by PCR and Western blot. Results The fbpA gene was not detected in genome of fbpA-deletion mutant of Listeria monocytogenes,and FbpA was not expressed in fbpA-deletion mutant of Listeria monocytogenes. Conclusion The fbpA-deletion mutant of Listeria monocytogenes was constructed successfully.%目的 构建单核细胞增生性李斯特菌fbpa基因敲除菌株.方法 克隆fbpa及其上、下游基因,构建其载体质粒；通过酶切反应将上、下游基因分别重组到载体质粒中,形成同源重组质粒；同源重组质粒电转入细菌内,进行同源重组；采用PCR、Western blot鉴定敲除菌株.结果 单核细胞增生性李斯特菌fbpa基因敲除菌株基因组DNA无fbpa基因片段,且无FbpA蛋白表达.结论 成功构建单核细胞增生性李斯特菌fbpa基因敲除菌株.