Gene Discovery and Expression Analysis using SAGE

使用 SAGE 进行基因发现和表达分析

• 批准号: 7304518
• 负责人: Denise Garcia
• 金额: $ 22.2万
• 依托单位: CALIFORNIA STATE UNIVERSITY SAN MARCOS
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-07-01 至 2011-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7304518
• 关键词: Aquaculture; Basic Science; Biological; Biological Process; Clinical; Complementary DNA; Condition; Count; Defense Mechanisms; Effectiveness; Expression Library; Future; Gene Expression; Generations; Genes; Genetic Transcription; Genome; Goals; Grant; Host Defense; IHHNV; Immune; Immune response; Immune system; Invertebrates; Knowledge; Libraries; Life; Methods; Microbe; Mind; Modeling; Molecular Profiling; Monitor; Organism; Pathologic; Pattern; Peptides; Process; Proteins; RNA; Reporting; Research; Sampling; Shrimp; Signal Transduction; Source; Statistical Methods; System; Techniques; Therapeutic; Time; Variant; Vertebrates; Viral; Viral Pathogenesis; Virus; Virus Diseases; White spot syndrome virus 1; antimicrobial peptide; base; cDNA Arrays; fluorophore; gene discovery; gene function; interest; microbial; novel; pathogen; research study; response; serial analysis of gene expression; tool

DESCRIPTION (provided by applicant): The primary goal of this project is to monitor global gene expression patterns in response to viral pathogenesis in the shrimp, Litopenaeus stylirostris. Changes in gene expression patterns have profound effects on biological functions. Increasingly, focused researched efforts are emerging to understand invertebrate defense systems and host response to pathogens. The innate immune strategies developed within invertebrate systems serve as models for understanding host defense strategies in response to viral pathogens for both vertebrates and invertebrates. To date, there are limited reports in penaeids using expression analysis to discover genes involved in the innate immune system and only one that has partially examined global gene expression. There are two techniques available for global gene expression analysis, cDNA microarray and serial analysis of gene expression (SAGE). Both are aimed at monitoring thousands of genes within a biological system under different experimental conditions, however, SAGE is the only technique that does not require prior knowledge of the genome of interest. In addition, SAGE analysis is based on the direct numerical counting of gene tags for comparison between libraries, rather than the arduous statistical methods required for microarray fluorophore signal intensity analysis. With this in mind, the objectives of this research are three-fold: 1) to compare global gene expression profiles between virally infected and healthy shrimp using SAGE, 2) to further characterize differentially expressed SAGE tags between healthy control (pathogen-free) versus virally infected shrimp using generation of longer cDNA fragments (GLGI) and 5' rapid amplification of cDNA ends (RACE), and 3) to determine expression levels of the differentially expressed genes for varying time points. The fulfillment of these objectives represents a unique opportunity to understand the invertebrate immune system as it relates to viral pathogenesis and will serve as a model to understand other animal, vertebrate and invertebrate, defense mechanisms. The primary goal of this project is to monitor global gene expression patterns in response to viral pathogenesis in the shrimp, Litopenaeus stylirostris. Increasingly, focused researched efforts are emerging to understand invertebrate defense systems and host response to pathogens. This project represents a unique opportunity to understand the invertebrate immune system as it relates to viral pathogenesis and will serve as a model to understand other animal, vertebrate and invertebrate, defense mechanisms.

描述（由申请人提供）：该项目的主要目标是监测细角对虾病毒发病机制的整体基因表达模式。基因表达模式的变化对生物功能具有深远的影响。越来越多的集中研究工作正在出现，以了解无脊椎动物的防御系统和宿主对病原体的反应。无脊椎动物系统内开发的先天免疫策略可作为了解脊椎动物和无脊椎动物针对病毒病原体的宿主防御策略的模型。迄今为止，对虾类使用表达分析来发现参与先天免疫系统的基因的报道有限，并且只有一份部分检查了整体基因表达。有两种技术可用于全局基因表达分析：cDNA 微阵列和基因表达系列分析 (SAGE)。两者都旨在在不同的实验条件下监测生物系统内的数千个基因，但是，SAGE 是唯一不需要事先了解感兴趣的基因组的技术。此外，SAGE 分析基于基因标签的直接数值计数以进行文库之间的比较，而不是微阵列荧光团信号强度分析所需的繁琐的统计方法。考虑到这一点，本研究的目标有三个方面：1) 使用 SAGE 比较病毒感染虾和健康虾之间的全局基因表达谱，2) 使用较长 cDNA 片段 (GLGI) 和 5' 快速扩增 cDNA 末端 (RACE) 的生成，进一步表征健康对照（无病原体）与病毒感染虾之间差异表达的 SAGE 标签，以及 3）确定差异表达基因的表达水平 时间点。这些目标的实现为了解无脊椎动物免疫系统提供了一个独特的机会，因为它与病毒发病机制有关，并将作为了解其他动物、脊椎动物和无脊椎动物防御机制的模型。该项目的主要目标是监测细角对虾病毒发病机制的整体基因表达模式。越来越多的集中研究工作正在出现，以了解无脊椎动物的防御系统和宿主对病原体的反应。该项目为了解与病毒发病机制相关的无脊椎动物免疫系统提供了一个独特的机会，并将作为了解其他动物、脊椎动物和无脊椎动物防御机制的模型。