Equipment for a molecular biology research facility

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This equipment grant is in support of my research on the genetic basis of stress-induced changes in virulence of enterhemorrhagic E. coli O157:H7 which is funded by an NSERC Discovery Grant (2006-2011).  This pathogen causes watery and bloody diarrhea and can lead to serious, even fatal complications including hemolytic uremic syndrome (HUS).  During ingestion, this pathogen is exposed to a variety of stresses including acid and bile salt stress as well as short chain fatty acid stress.  Our research has shown that acid and bile salt stress enhance the ability of this organism to adhere to host epithelial cells and we are currently examining the genetic basis of this enhanced adhesion.  We are also assessing the effect of short chain fatty acid stress on bacterial-host adhesion as well as the impact of these ingestion stresses on the organism's ability to produce verotoxin, a potent cytotoxin highly associated with the development of HUS. Molecular biology tools are being used to determine the genetic profile of this organism before and after each of the stresses and to identify molecular candidates that play a role in the altered virulence after stress. Once identified, we will use mutational analysis to assess the role of each of these candidates in virulence assays.  The Nanodrop spectrophotometer is necessary to quantify microvolumes of bacterial DNA, RNA and proteins. Microarray analysis equipment will be used to determine genetic expression profiles of the pathogen. The microfuge/centrifuge is critical to all aspects of work with DNA, RNA, bacterial and cell infection work.  Without this equipment, my students must travel to other labs in Ryerson and even in other institutions to use similar equipment.  The purchase of this equipment is very important to my research program and to those of the coapplicants and will directly contribute to the training of young Canadian scientists including graduate and undergraduate students as well as postgraduate research assistants (over 35 total for all applicants). My research program has significant potential benefits for the development of therapeutic treatments of this infection through the identification of new virulence properties and molecular targets for antimicrobial agents.