Acquisition of DIGE Instrumentation to Enhance Proteomic Research in Nevada

描述（由申请人提供）：本提案是2007年3月最初提交的NCRR共享仪器赠款的重新提交。内华达州蛋白质组学中心与INBRE和COBRE项目以及NIH资助的研究人员合作，要求提供设备和软件，使我们的2-D凝胶实验室能够进行差异凝胶电泳（DIGE）实验。请求提供的物品包括：Typhoon Trio激光成像仪、DeCyder v6.5和DeCyder扩展数据分析软件以及带有DIGE升级的Genomics Solution ProPic II Spot Cutter。二维（2-D）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2-D凝胶是出了名的不一致，并且凝胶间的变化使得难以准确测定蛋白质丰度的变化。DIGE技术克服了这些障碍，运行控制和实验样品，沿着一个额外的归一化样品，在同一个2-D凝胶。这3种类型的样品中的每一种都用不同的荧光染料标记;这些染料通过Typhoon成像仪检测，染料强度的差异通过DeCyder软件定量。通过ProPic II斑点切割器从凝胶上切割目标蛋白质斑点，并通过质谱法进行鉴定。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良好。我们的2-D凝胶实验室是由BRIN和EPSCoR资助建立的，拥有5年的经验以及生产凝胶的所有必要设备。DIGE技术对于我们在内华达州大学的许多NIH资助的研究人员的研究将是非常宝贵的，他们试图检测各种疾病状态中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制成的蛋白质在细胞之间差异很大，并且会因环境影响或疾病状态而变化。通过研究细胞中蛋白质的数量变化或化学修饰，我们可以更好地了解疾病的分子基础以及如何治疗它。在这项资助计划中，我们要求资助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.