Quorum sensing regulation of bacterial development

项目摘要 群体感应是一种细胞间通讯的方法，被细菌用来检测和 应对人口密度的变化，控制有利于群体的行为， 在其环境中的适应和生存（例如，共生和发病机制）。我的研究 该计划的重点是细菌如何使用群体感应信号来控制表达， 毒力基因为了实现这一目标，我们研究了弧菌属物种中的群体感应基因调控， 既作为相关病原体，又作为已建立的群体感应模型系统。我们专注于 主弧菌群体感应转录因子LuxR，其在所有弧菌中保守 是数百个基因的核心调节器。我的研究小组利用微生物 遗传学来剖析影响基因的弧菌物种的LuxR调控网络， 表达毒力。我的实验室已经为每一个建立了表型实验 途径，包括III型分泌，蛋白酶产生，生物膜形成，生物发光， 渗透胁迫反应和自然转化。这些实验是 我的研究计划的组成部分。 我申请资金购买BioTek Synergy H1混合多模式 单色荧光酶标仪。这个酶标仪测量 吸光度、荧光和生物发光，在宽范围内具有可调精度， 在可调节的温度孵育和振荡参数下，我们经常 进行生长、生物发光产生、荧光和lacZ报告基因的测定， 测量基因表达、表型分析、酶动力学和定量 核酸和蛋白质的测量。对于这些测定中的每一种，我们都严重依赖于 酶标仪的高通量性质，可在96孔或384孔中进行实验 在平板上频繁读数24小时。我们目前正在使用BioTek Cytation 3是我的实验室和其他三个相邻实验室经常使用的。24小时 我们进行的生长/生物发光测定严重限制了我们进行 额外的分析。购买Synergy H1酶标仪将使我的实验室能够 增加实验输出，并将此酶标仪专用于我们实验室，优先考虑24- 小时生长/荧光测定。

Project summary Quorum sensing is a method of cell-cell communication used by bacteria to detect and respond to changes in population density and control behaviors that benefit the group for adaption and survival in their environment (e.g., symbiosis and pathogenesis). My research program is focused on how bacteria use quorum sensing signaling to control expression of virulence genes. Toward this goal, we study quorum sensing gene regulation in Vibrio species, both as relevant pathogens and as established quorum sensing model systems. We focus on the master Vibrio quorum sensing transcription factor LuxR, which is conserved in all Vibrio species and is the core regulator of hundreds of genes. My research group uses microbial genetics to dissect the LuxR regulatory networks of Vibrio species that influence gene expression for virulence. My lab has established phenotypic experiments for each of these pathways, including type III secretion, protease production, biofilm formation, bioluminescence, osmotic stress response, and natural transformation. These experiments are the foundational components of my research program. I am requesting funds to purchase a BioTek Synergy H1 Hybrid Multi-Mode Monochromator Fluorescence Microplate Reader. This microplate reader measures absorbance, fluorescence, and bioluminescence with tunable precision across a wide range of wavelengths with both adjustable temperature incubation and shaking parameters. We routinely perform assays for growth, bioluminescence production, fluorescence and lacZ reporters for measuring gene expression, phenotypic assays, enzyme kinetics, and quantitation measurements of nucleic acids and proteins. For each of these assays, we heavily rely on the high-throughput nature of the microplate reader to conduct experiments in 96-well or 384-well plates with readings at frequent intervals for 24 hours. We are currently using a BioTek Cytation3 that is used constantly by my lab and three other neighboring labs. The 24-hour growth/bioluminescence assays that we perform are severely limiting our time to perform additional assays. The purchase of the Synergy H1 microplate reader will enable my lab to increase experimental output and dedicate this plate reader solely to our lab and prioritize to 24- hour growth/fluorescence assays.