Construction of the recombinant strains of Listeria monocytogenes expressing green fluorescent protein and characterization of their biological properties
Abstract：To evaluate the immune responses induced by transmission of the exogenous antigens through Listeria monocytogenes in the study of prokaryotic transmission model of green fluorescent protein(GEP),the promoter of listeria lysine O(LLO) and the GFP gene gfp were fused together by using the SOEing PCR and then integrated to yzuLM1-2 actA and plcB gene fragments through homologous recombination.In this way,GFP was expressed through the function of up-stream LLO promoter.It was verified by PCR amplification that gfp gene could be successfully integrated to listerial genome.The result in animal experiment showed that the LD50 of the recombinant bacteria in mice was 4.31×108,that was significantly reduced to 10 folds in comparison with that of the attenuated strain yzuLM1-2.The sera of mice immunized with recombinant bacteria showed specific band at 27 kDa in Western blot assay and could elicit production of high level of anti-GFP antibodies as demonstrated by ELISA assay.It is evident that the GFP transmitted by the attenuated L.monocytogeneses can induce strong immunogenicity in mice,thus constituting a solid basis to elucidate the molecular mechanism of pathogenesis and immunity triggered by L.moncytogenes.
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