[Construction of luxS gene knockout mutant of Enterococcus faecalis].

ABSTRACT

PUPPOSE To construct quorum sensing luxS knockout mutants of Enterococcus faecalis through homologous recombination. METHODS: The upstream and downstream flank DNA fragments of E. faecalis luxS gene (up, dn) and erythromycin resistance gene (erm) were amplified by PCR. In order to construct recombination plasmid Puemrd, these DNA fragments were inserted into the plasmid pUC18 by corresponding double digests. After allelic exchange, the luxS knockout mutants strains were selected on 30 µg/mL erythromycin plates. RESULTS: With endonuclease reaction and DNA sequencing, it was proved that the objective plasmid, Puemrd, was constructed correctly. The luxS knockout mutants strains were confirmed by PCR. CONCLUSIONS: Enterococcus faecalis luxS gene has been successfully disrupted with homologous recombination. This mutant strain sets a good foundation for further functional study.