Acquisition of DIGE Instrumentation to Enhance Proteomic Research in Nevada

描述（由申请人提供）：本提案是对最初于2007年3月提交的NCRR共享仪器拨款的重新提交。内华达蛋白质组学中心与INBRE和COBRE项目以及美国国立卫生研究院资助的研究人员合作，要求设备和软件，使我们的二维凝胶实验室能够进行差异凝胶电泳（DIGE）实验。所要求的项目包括：台风Trio激光成象仪，DeCyder v 6.5和DeCyder扩展数据分析软件，以及带有DIGE升级的Genomics Solution proic II点切割机。二维凝胶是目前蛋白质组学实验中分离和定量蛋白质的主要方法。然而，标准的二维凝胶是出了名的不一致，凝胶与凝胶之间的差异使得准确测定蛋白质丰度的变化变得困难。DIGE技术通过在相同的二维凝胶上运行对照和实验样品以及额外的正态化样品来克服这些障碍。这三种样品中的每一种都用不同的荧光染料标记；这些染料由台风成像仪检测，染料强度的差异由DeCyder软件量化。用proic II点切割器从凝胶上切下感兴趣的蛋白点，并用质谱法进行鉴定。所要求的系统代表了最先进的DIGE技术，并与我们现有的仪器集成得很好。我们的二维凝胶实验室是由BRIN和EPSCoR资助建立的，拥有5年的经验和生产凝胶所需的所有设备。DIGE技术对于我们在内华达大学的许多美国国立卫生研究院资助的研究人员的研究是无价的，他们试图检测各种疾病状态下生理上重要的蛋白质变化。公共卫生相关性：虽然任何一种生物体中的所有细胞都含有相同的DNA，但由这种DNA制成的蛋白质在不同细胞之间差异很大，并且随着环境影响或疾病状态的变化而变化。通过研究细胞中蛋白质数量的变化或化学修饰，我们可以更好地了解疾病的分子基础以及如何治疗疾病。在这项拨款申请中，我们要求资金购买一种敏感和准确的方法来比较健康和疾病状态下细胞中产生的蛋白质数量。

DESCRIPTION (provided by applicant): This proposal is a resubmission of an NCRR Shared Instrumentation Grant originally submitted in March 2007. The Nevada Proteomics Center, in collaboration with INBRE and COBRE projects and NIH-funded investigators, requests equipment and software that will enable our 2-D Gel Laboratory to perform differential in-gel electrophoresis (DIGE) experiments. The requested items include: a Typhoon Trio laser imager, DeCyder v 6.5 and DeCyder Extended Data Analysis software, and a Genomics Solution ProPic II Spot Cutter with DIGE Upgrade. Two-dimensional (2-D) gels are currently the principle method for separating and quantifying proteins in proteomics experiments. However, standard 2-D gels are notoriously inconsistent and gel-to-gel variations make accurate determinations of change in protein abundance difficult. DIGE technology overcomes these obstacles by running a control and experimental sample, along with an additional normalizing sample, on the same 2-D gel. Each of these 3 types of samples is labeled with a different fluorescent dye; these dyes are detected by the Typhoon imager and differences in dye intensities are quantified by DeCyder software. Protein spots of interest will be cut from the gel by the ProPic II spot cutter and identified by mass spectrometry. The requested system represents the state-of-the-art in DIGE technology and integrates well with our existing instrumentation. Our 2-D Gel Lab, which was set up with BRIN and EPSCoR funding, has 5 years of experience plus all necessary equipment for generating the gels. The DIGE technology would be invaluable for the research of many of our NIH-funded investigators at University of Nevada who are attempting to detect physiologically important protein changes in a variety of disease states. PUBLIC HEALTH RELEVANCE: While all cells in any one organism contain the same DNA, the proteins that are made from this DNA vary greatly from one cell to another and vary in response to environmental influences or disease states. By studying changes in the amounts or chemical modifications of proteins in a cell, we can better understand the molecular basis of a disease and how to treat it. In this grant proposal, we request funds to purchase a sensitive and accurate way to compare the amounts of proteins produced in a cell under healthy and disease states.