Abstract: 【Objective】 The experiment was aimed to construct a strain of Listeria monocytogenes with gltC gene deletion and elucidate the effect of gltC gene on the acid stress resistance of Listeria monocytogenes.【Method】 The effect of glutamic acid on the survival ability of the 10403S was determined by acid stress test with or without the addition of exogenous glutamic acid under lethal acidic conditions.The homologous recombination method was used to construct a strain with a deletion of gltC gene.The effect of gltC gene deletion on the growth capacity of the strain was determined by growth curves.The effect of gltC gene deletion on the acid stress capacity of Listeria monocytogenes was determined by the acid stress survival assay.The effect of the gltC gene deletion on the expression level of GadD2 protein was determined by Western blotting.By constructing the gfp reporter plasmid carrying the gltBD promoter region,the effect of gltC gene deletion on the transcription level of the gltBD promoter region was measured under acidic and neutral conditions.【Result】 Acid stress results showed that adding exogenous glutamic acid could extremely significantly improve the survival ability of strain 10403S under acidic conditions at pH 2.5 (P＜0.01).PCR results showed that the deletion strain 10403SΔgltC was successfully constructed.The growth curve results showed that gltC gene deletion did not affect the growth ability of Listeria monocytogenes.The results of acid stress showed that the growth ability of the deletion strain 10403SΔgltC was weaker than that of the parental strain 10403S in a lethal acidic environment at pH 2.5 (P＜0.01).The results of the Western blotting showed that the deletion of gltC gene did not affect the expression level of GadD2 protein (P＞0.05).PCR results showed that gfp with gltBD promoter region reported plasmid was successfully constructed.GFP fluorescence analysis showed that the deletion of gltC gene extremely significantly decreased the transcription level of the gltBD promoter region under both acidic and neutral conditions (P＜0.01).【Conclusion】 Deletion of gltC gene did not affect the normal growth of the Listeria monocytogenes 10403S,but the deletion strain 10403SΔgltC reduced resistance to acid stress and decreased the level of transcription in the gltBD promoter region,without affecting the level of GadD2 protein expression.

Key words: Listeria monocytogenes; glutamate synthase regulatory factor; gltC gene; acid stress resistance; reporter gene