Equipment for a molecular biology research facility

这项设备资助是为了支持我对肠出血性大肠杆菌O157:H7的应激诱导毒力变化的遗传基础的研究，该研究由NSERC发现资助（2006-2011）。这种病原体引起水样和带血腹泻，并可导致严重甚至致命的并发症，包括溶血性尿毒症综合征（HUS）。在摄入过程中，这种病原体暴露于多种应激，包括酸和胆盐应激以及短链脂肪酸应激。我们的研究表明，酸和胆盐胁迫增强了这种生物体粘附宿主上皮细胞的能力，我们目前正在研究这种增强粘附的遗传基础。我们还评估了短链脂肪酸应激对细菌-宿主粘附的影响，以及这些摄入应激对生物体产生维罗毒素的能力的影响，维罗毒素是一种与溶血性尿毒综合征的发展高度相关的强效细胞毒素。分子生物学工具被用来确定这种生物体在每次应激前后的遗传谱，并确定在应激后毒性改变中起作用的候选分子。一旦确定，我们将使用突变分析来评估这些候选物在毒力分析中的作用。纳米滴分光光度计是定量细菌DNA、RNA和蛋白质微体积所必需的。微阵列分析设备将用于确定病原体的基因表达谱。在DNA、RNA、细菌和细胞感染工作的各个方面，微生物/离心机都是至关重要的。如果没有这些设备，我的学生就必须到瑞尔森的其他实验室甚至其他机构去使用类似的设备。这台设备的购买对我的研究计划和共同申请者的研究计划非常重要，并将直接有助于培养年轻的加拿大科学家，包括研究生和本科生以及研究生研究助理（所有申请人总数超过35人）。我的研究项目通过鉴定抗菌剂的新毒力特性和分子靶点，对这种感染的治疗方法的发展具有重大的潜在益处。

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.