The Screening Of Listeria Monocytogenes Mutants On Biofilm Formation

Listeria monocytogenes is one of the most important foodborn pathogens. Characterization of the functional genes and related regulation elements is the basis to understand the mechanism of bacterial biofilm formation, which is important for the development of prevention methods to the bacterial pollution. The transposon Tn917 was used to produce insertional mutagenesis of L. monocytogenes by electroporation and transposition, and 1880 mutants were obtained in this study, which extended the mutants in our lab to the total amounts of 2200. A thousand of mutants was screened for the negative mutation on biofilm formation by crystal violet staining analysis after microtiter incubation. Eight strains with poor ability to form biofilm were finally sorted out, and the rate of targeted mutation amounts was estimated as 8‰. The poor ability on biofilm formation of these mutants was further confirmed by fluorescence microscopy. It was also confirmed by PCR assay that the mutation was generated by the insertion of Tn917 into the genomes of those targeted mutants.The genomic DNA of mutant LM-301 was involved in the analysis of IPCR ( inverse PCR), which indicates the interrupted gene segment. The DNA sequencing and GenBank alignment results showed the gene encoded the cell wall surface anchor family protein.