Study of cellular location preferences of Salmonella typhimurium along with gene expression analysis

Salmonella enterica serovar typhimurium a hyperreplicating population, exists inside the cytoplasm of infected epithelial cells. This population expresses genes belonging to the Salmonella Pathogenicity Island 1 (SPI-1) after exiting from the Salmonella Containing Vacuole (SCV) (Knodler, et al., 2010) (Beuzon, et al., 2002). SCV provides a protective niche to Salmonella typhimurium and the reasons for its escape into the cytoplasm are not known. We wanted to isolate cytoplasmic as well as SCV contained Salmonella typhimurium, so as to understand their cellular location preferences and to study the gene expression and regulation. In order to achieve this, we wanted to make use of the fact that vacuolar bacteria have their Salmonella Pathogenicity Island 2 (SPI-2) genes turned on, while those that exit from the vacuole switch off this system and express SPI-1 genes(Knodler, et al., 2010). Thus, a time based expression tracking of SPI-1 and SPI-2 in epithelial Salmonella typhimurium would help us to determine their cellular location and using fluorescent reporters as trackers they would be separated with the help of FACS (Fluorescence Activated Cell Sorting). This gave rise to the objective of my work: Creation of a molecular tool to study temporal gene expression in bacteria. Keywords— SCVSalmonella Containing Vacuole, T3SSType Three Secretory System, TspTail Specific Potease, SPISalmonella Pathogenicity Island, RFPRed Fluorescent Protein, HBI 4 (phydroxybenzylidene ) imidazolidin-5-one