Module 3: Gene Expression/Proteomics

基因表达和蛋白质组学模块将为研究人员提供设计和 进行与视力疾病相关的全球基因表达谱和蛋白质组改变的研究 系统。在过去的二十年里，已经开发了许多高吞吐量平台来研究 全球基因表达模式。这些技术包括斑点cDNA或寡核苷酸阵列、Affymetrix 基因芯片微阵列、Illumina珠芯片微阵列和最新的下一代测序 技术包括RNA测序(RNA-Seq)和单细胞RNA测序(scRNA-seq)。此核心 将使用RNA-Seq和scRNA-seq进行基因表达研究。同样，液体的发展-- 在过去的十年中，色谱质谱(LC-MS/MS)使蛋白质组学分析发生了革命性的变化 通过其前所未有的深度、速度和准确性，即使在小体积样品的情况下也是如此。最新的一款 这项技术的突破是Orbitrap Fusion三元质谱仪，它允许研究人员 扩大蛋白质组学分析的范围，包括翻译后修饰。我们有这台乐器在 奥古斯塔大学和最近，我们成功地建立了一个分析水溶液的协议 使用这个设备的幽默蛋白质组。这些最新的测序和质谱仪技术，用于评估基因 表达和蛋白质水平，可能会让研究人员望而却步，他们的科学专长在其他地方。这个 新技术也带来了与高通量数据分析和解释相关的挑战。至 绕过这些障碍，基因表达和蛋白质组学模块旨在引导研究人员 实验设计、样品制备、适当技术的深思熟虑的选择和生物信息学 支持测序和蛋白质组学分析。此外，该核心还将帮助研究人员 解释结果并设计进一步的实验以确认/确认研究结果。的目标是 本模块用于指导视觉研究人员进行高通量的基因表达分析(使用测序) 以及眼组织的蛋白质组学分析(使用质谱学)。本模块中提供的服务反映了 敏锐地意识到与特殊眼组织实验相关的细微差别，并将 在两个目标上实现。目标1：为核心调查人员提供专家咨询，以分析 可视化系统，关于实验设计、样品制备和基因传导技术的选择 眼组织的表达研究和/或蛋白质组学分析。目标2：为核心调查人员提供优秀的 生物信息学支持分析高通量基因后产生的大数据集 眼组织的表达或蛋白质组学分析。

The gene expression and proteomics module will provide investigators with expertise in designing and performing studies of global gene expression profiles and proteomic alterations related to diseases of the visual system. In the last two decades, a number of high throughput platforms have been developed to investigate global gene expression patterns. These technologies include spotted cDNA or oligonucleotide arrays, Affymetrix GeneChip microarrays, Illumina BeadChip microarrays, and most recently next-generation sequencing technologies including RNA sequencing (RNA-Seq) and Single-cell RNA sequencing (scRNA-seq). This Core will use RNA-Seq and scRNA-seq for gene expression studies. Similarly, the development of Liquid- Chromatography Mass Spectrometry (LC-MS/MS) over the last decade has revolutionized proteomic analyses through its unprecedented depth, speed and accuracy even with small volume samples. One of the latest breakthroughs in this technology is the Orbitrap Fusion Tribrid mass spectrometer, which allows researchers to extend the limits of proteomic analyses including post-translational modifications. We have this instrument at Augusta University and recently, we have successfully established a protocol for the analysis of the aqueous humor proteome using this device. These latest sequencing and mass-spec technologies, used to evaluate gene expression and protein levels, can be intimidating to researchers, whose scientific expertise lies elsewhere. The new technologies also present challenges associated with high throughput data analyses and interpretation. To circumvent these hurdles, the gene expression and proteomics module is intended to guide researchers in experimental design, sample preparation, thoughtful selection of appropriate technology, and bioinformatics support for sequencing and proteomic profiling. In addition, the core will also help researchers with the interpretation of results and designing further experiments for confirmation/validation of findings. The goal of this module is to guide vision researchers with high throughput gene expression analyses (using sequencing) and proteomic analyses (using mass-spectrometry) of ocular tissues. The services offered in this module reflect keen awareness of the nuances related to experimentation with specialized ocular tissues and will be accomplished in two aims. Aim 1: Provide core investigators with expert consultation, tailored to analysis of the visual system, on experimental design, sample preparation and selection of technology to conduct gene expression studies and/or proteomic analyses of ocular tissues. Aim 2: Provide core investigators with excellent bioinformatics support in analyzing the large data sets that are generated following high throughput gene expression or proteomic analyses of ocular tissues.