Protein Expression and Proteomics

蛋白质表达和蛋白质组学(PEP)核心从一年开始获得补充资金 2006年6月1日，为我们的调查人员社区提供一个未得到满足的需求。该蛋白质表达部分 在约翰·埃尔德博士的指导下，拟议的试点核心将提供适当的载体和 细胞系需要促进研究人员靶向蛋白的表达克隆。的蛋白质组学部分 在Bruce Torbett博士的指导下，核心将为CFAR提供蛋白质分析服务 调查人员。这些服务包括从病毒、细胞和组织中提取蛋白质；蛋白质解析 使用一维和二维(D)电泳法和成像系统进行定量；蛋白质 使用质谱学进行鉴定；使用Luminex？xMAP？多重分析蛋白质 检测技术。PEP核心的独特之处在于专注于为调查人员提供服务 要求表达HIV-1和其他病毒基因的蛋白质以及HIV-1和其他病毒的蛋白质分析 病毒感染的样本。 核心的蛋白质表达部分将提供所需的适当载体和细胞系 使用细菌、昆虫细胞或哺乳动物细胞促进研究人员靶向蛋白的表达克隆 被认为适合特定应用的矢量系统。Elder实验室编制了一份 为每个表达系统提供了大量的载体，并在蛋白质表达方面拥有丰富的经验 以及用于配体结合研究、酶学和结晶学分析的提纯。实验室是 配备在所需温度下培养细胞所需的孵化器和离心机，FPLC， 和高效液相色谱系统，在必要时帮助进行蛋白质纯化。核心将协助调查员 载体的选择及靶基因的扩增和克隆。此外，核心将帮助调查人员 在优化蛋白质表达和选择最适合每个蛋白质的纯化方案方面 申请。 核心的蛋白质组学部分将提供蛋白质解析的所有方面，从可视化到 身份证明。CFAR成员将可以访问NCI资助的蛋白质组学核心 斯克里普斯研究所分子和实验医学系。目前， 蛋白质组学核心公司拥有可用的一维和二维凝胶电泳蛋白质分离仪器，IEF 用于溶液相IEF的变焦分级器，一种最先进的具有Bio-Rad的二维蛋白质组成像系统 VersaDoc 1000成像仪和成像软件。所有成像软件均可免费使用 安装在核心内的常驻计算机。核心技术人员提供技术建议，以及 需要时，核心技术人员会提供有关分析和成像软件的演示和研讨会。在……里面 此外，核心技术人员将为调查人员提供优化蛋白质提取程序的建议 以及选择最适合每种应用的纯化方案。 蛋白质表达和蛋白质组学核心将提供关于核心的信息研讨会/会议 每年提供服务，并在整个供资期间举办“新技术”研讨会/会议。

The Protein Expression and Proteomics (PEP) Core received supplemental funding for one year starting June 1, 2006, to provide an unmet need for our community of investigators. The Protein Expression portion of the proposed pilot Core, under the direction of Dr. John Elder, will provide the appropriate vectors and cell lines needed to facilitate expression cloning of investigator-targeted proteins. The Proteomics portion of the core, under the direction of Dr. Bruce Torbett, will provide protein analysis services for CFAR investigators. These services include protein extraction from viruses, cells, and tissues; protein resolution and quantification using 1- and 2-dimensional (D) electrophoresis and imaging systems; protein identification using mass spectroscopy; and analyses of proteins using Luminex¿ xMAP¿ multiplex detection technology. The unique aspect of the PEP Core is the focus on providing services to investigators requiring expression of proteins from HIV-1 and other viral genes and protein analysis of HIV-1 and other virally infected samples. The Protein Expression portion of the Core will provide the appropriate vectors and cell lines needed to facilitate expression cloning of investigator-targeted proteins, using bacterial, insect cell, or mammalian cell vector systems, as deemed appropriate for the particular application. The Elder laboratory has compiled an extensive panel of vectors for each expression system and has extensive experience in protein expression and purification for ligand binding studies, enzymology, and crystallographic analyses. The laboratory is equipped with incubators required to grow cells at the required temperatures and also centrifuges, FPLC, and HPLC systems to aid in protein purification where necessary. The Core will aid the investigator in vector selection and amplification and cloning of targeted genes. In addition, the Core will help investigators in optimizing protein expression and selection of purification schemes most appropriate for each application. The Proteomics portion of the Core will provide all facets of protein resolution from visualization to identification. CFAR members will have access to the NCI-funded Proteomics Core housed within the Departmental of Molecular and Experimental Medicine at The Scripps Research Institute. Currently the Proteomics Core has available 1- and 2-D gel electrophoresis protein separation instrumentation, IEF Zoom fractionators for solution phase IEF, a state-of-the-art 2D Proteomic Imaging System with a Bio-Rad VersaDoc 1000 imager and imaging software. All imaging software can be utilized free of charge on resident computers housed within the Core. Technical advice is available from the Core technicians, and when needed, Core technicians provide demonstrations and seminars on analysis and imaging software. In addition, Core technicians will provide advice to investigators for optimizing protein extraction procedures and selection of purification schemes most appropriate for each application. The Protein Expression and Proteomics Core will provide informational seminars/sessions on the Core services each year and "new technology" seminars/sessions throughout the funding period.