In this study, we aimed at identifying the regulatory role ofmarTgene, known as the regulator ofmisL, on 15 different biofilm-related genes inS.Typhimurium 14028 strain. We also tested the strains for their ability to form biofilm and determined the adherence characteristics of the wild type and the mutant strains of the organism on Caco-2 and HEp-2 cells. For comparative analyses of the candidate genes, individual gene mutations were created via antibiotic gene cassette insertion into each gene of interest.marTgene was cloned behind an arabinose inducible BAD promoter in order to controlmarTexpression. This recombinant plasmid was transfer into each of the 15 mutant strains to investigate the level of expression of each single gene in the presence and absence ofmarTinduction. Besides determination of variations in biofilm formation by each mutant strain, the attachment characteristics of them onto Caco-2 and HEp-2 cell lines were also reported. As a result of attachments experiments on polystyrene surfaces, it was determined that the biofilm production capacity of each mutant strain decreased in a statistically significant manner (p < 0.05). QRT-PCR trials indicated that themarTgene regulates the expression of 14 genes, namelyfimA,fimD,fimF,fimH,stjB,stjC,csgA,csgD,ompC,sthB,sthE,rmbA,fliZandyaiC, in a positive manner. QRT-PCR studies were also revealed that the MarT protein positively regulates its own promoter. When the adherence characteristics of the mutant strains and the wild-type were investigated by using Caco-2 and HEp-2 cells, it was determined that the single gene mutations did have no effect on bacterial adhesion. In view of our mutational analyses and biofilm formation studies, it was concluded thatfliZ,ompC,rmbA,stjBandstjCgenes are related with biofilm formation inSalmonella,besides other cellular functions of them. Taken together, our data suggested that the regulatory role of MarT protein is not only restricted to the regulation ofmisLgene expression, but it rather acts as a general regulator on the biofilm-related genes inSalmonella.