Core--Analysis

描述(由申请人提供)： 分析核心将提供与基因治疗相关的分子分析方法以及细胞和组织对基因转移的反应的专业知识、人员和设备。分析核心位于新装修的Belfer基因治疗核心设施中，由实验室空间和包含序列检测系统7700(TaqMan)实时定量聚合酶链式反应系统和BIOMEK FX样品准备机器人的设备室组成。该核心将向该计划项目的研究人员提供几种与基因治疗相关的先进分子技术。其中包括：自动提取和纯化DNA；用于DNA定量的TaqMan定量聚合酶链式反应和用于评估基因表达水平变化的定量RT-PCR。高通量基因分型、基因表达微阵列(Affymetrix GeneChip)以及基因组和微阵列表达数据的分析也是可用的，尽管不是该计划项目中拟议的研究的一部分。所有四个项目都将平等地使用分析核心，主要是在定量聚合酶链式反应领域，用于定量细胞和组织中的载体DNA，以及在定量RT聚合酶链式反应领域，用于基因表达水平的绝对和相对量化。分析核心主任尼尔·哈克特花了相当长的时间优化定量聚合酶链式反应(TaqMan)方法来评估基因转移，并展示了该方法的潜在威力和高通量 正确执行。由于分子技术的复杂性和成本以及所需的先进设备，对于任何一个小组来说，获得通过分析核心获得的所有复杂的方法和通过高通量获得的效率都是一个挑战。

DESCRIPTION (provided by applicant): The Analysis Core will provide expertise, personnel and equipment for molecular analysis methods relevant to gene therapy and the response of cells and tissues to gene transfer. The Analysis Core is located in the newly renovated Belfer Gene Therapy Core Facility and consists of laboratory space and an equipment room containing the Sequence Detection System 7700 (TaqMan) realtime quantitative PCR system and BioMek FX sample preparation robot. The Core will make available to the investigators of the Program Project several advanced molecular techniques relevant to gene therapy. These include: automated DNA extraction and purification; TaqMan quantitative PCR for DNA quantitation and quantitative RT-PCR for assessment of changes in gene expression levels. Also available, although not specifically part of the proposed studies in the Program Project, are high throughput genotyping, gene expression microarrays (Affymetrix GeneChip), and analysis of genomic and microarray expression data. All four projects will make equal use of the Analysis Core, primarily in the area of quantitative PCR for quantitation of vector DNA in cells and tissues and in quantitative RTPCR for the absolute and relative quantitation of gene expression levels. The Director of the Analysis Core, Neil Hackett, has spent considerable time optimizing quantitative PCR (TaqMan) methods to assess gene transfer and demonstrated the potential power and high throughput of the method when properly executed. Due to the complexity and expense of molecular techniques and the advanced equipment required, it is a challenge for any one group to acquire all of the sophisticated methodologies and efficiency derived from high throughput that are available through the Analysis Core.