Proteomics Core

蛋白质组学核心（核心B）提供了最先进的仪器和专业知识， 蛋白质组学的研究人员在这个PPG。蛋白质组学核心的一个独特功能是所有的可用性 主要的蛋白质组学分析技术，在分析方法上提供了卓越的灵活性。的 Proteomics Core提供1）简单和复杂多蛋白样品中的蛋白质鉴定， 包括多蛋白复合物的蛋白质成分，2）差异蛋白质的定量分析 表达和修饰，以及3）蛋白质修饰的分析。 蛋白质组学核心人员提供实验设计和样品制备方面的咨询， 消化、蛋白质和肽分离、MS分析和主要数据处理。两种电喷雾 用于LC-MS-MS实验的离子化（ESI）质谱仪和MALDI TOF-TOF仪器是 可用于MS分析。DIGE/MS技术包括荧光染料和多重2D凝胶， 用于直接定量比较的内部标准。自动化工作站用于蛋白质切除 和样品处理，然后进行MALDI-TOF MS和TOF/TOF MS/MS和自动数据库 用Mascot软件进行蛋白质鉴定。Core还提供了使用Nterminal的相对定量 反应性差异标记试剂（iTRAQ和同位素标记的苯基异氰酸酯），用于相对 修饰的蛋白质形式的定量。对于多蛋白复合物的蛋白质鉴定， 多维LC直接与自动化LC-MS-MS耦合，然后自动化数据库 用Sequest和Mascot软件搜索序列搜索是在高速多处理器上完成的 研究和教育高级计算中心（ACCRE）中的Linux集群。基于封存 使用开源工具（肽/蛋白质先知）对鉴定进行评估和过滤， 使用定制设计的数据库系统（CHIPS）来组织、过滤和比较数据集。 通过SALSA和P-Mod的额外数据分析能够鉴定翻译后修饰， 加合物或序列变体。 项目1（Peek）将利用DIGE/MS差分显示技术，项目3（Cover）将 从简单和复杂的多蛋白质样品中利用蛋白质鉴定技术。计划2 （Polk）将大量使用这两种服务。在修订后的《生物多样性公约》中不再提出蛋白质修饰分析。 项目，但可用于后续/补充研究。

The Proteomics Core (Core B) provides state-of-the-art instrumentation and expertise in analytical proteomics to investigators in this PPG. A unique feature of the Proteomics Core is the availability of all major proteomics analysis technologies, which offers exceptional flexibility in analytical approaches. The Proteomics Core provides 1) identification of proteins in both simple and complex multiprotein samples, ncluding protein components of multiprotein complexes, 2) quantitative analyses of differential protein expression and modification, and 3) analyses of protein modifications. Proteomics Core staff provide consultation on experimental design and sample preparation, as well as digestion, protein and peptide separations, MS analyses and primary data workup. Both electrospray onization (ESI) mass spectrometers for LC-MS-MS experiments and MALDI TOF-TOF instruments are available for MS analyses. DIGE/MS technology includes fluorescent dyes and multiplexed 2D gels with nternal standards for direct quantitative comparisons. Automated workstations are used for protein excision and sample processing, followed by MALDI-TOF MS and TOF/TOF MS/MS and automated database searching with Mascot software for protein identification. The Core also offers relative quantitation with Nterminal reactive differential labeling reagents (iTRAQ and isotopically-labeled phenylisocyanate) for relative quantitation of modified protein forms. For protein identification from multiprotein complexes, multidimensional LC is coupled directly with automated LC-MS-MS, followed by automated database searching with Sequest and Mascot software. Sequest searches are done on a high speed, multiprocessor Linux cluster in the Advanced Computing Center for Research and Education (ACCRE). Sequest-based identifications are evaluated and filtered using open-source tools (Peptide/Protein Prophet) and integrated with a custom-designed database system (CHIPS) for organizing, filtering and comparing data sets. Additional data analysis by SALSA and P-Mod enables identification of posttranslational modifications, adducts or sequence variants from MS-MS data. Project 1 (Peek) will utilize the DIGE/MS differential-display technology, and Project 3 (Cover) will utilize the protein identification technologies from both simple and complex multiprotein samples. Project 2 (Polk) will heavily utilize both services. Analysis of protein modifications is no longer proposed in the revised Projects, but may be used for subsequent/complementary studies.