Optimizations of rRNA-based Microbial Analyses

基于 rRNA 的微生物分析的优化

• 批准号: 7622594
• 负责人: Gary J. OLSEN
• 金额: $ 7.93万
• 依托单位: UNIVERSITY OF ILLINOIS AT URBANA-CHAMPAIGN
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-07-01 至 2011-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7622594
• 关键词: Address; Attention; Binding; Binding Sites; Biology; Clinical; Cloning; Computer Analysis; DNA; Data; Databases; Detection; Disease; Ecology; Ensure; Environment; Gene Amplification; Gene Targeting; Genes; Genome; Genomics; Goals; Growth; Health; Human; Illinois; Institutes; Kinetics; Medical; Membrane; Metagenomics; Methods; Microbe; Molecular; Nucleotides; Organism; Phase; Physiological; Play; Polymerase Chain Reaction; Population; Process; Protocols documentation; RNA Sequences; Relative (related person); Research; Research Personnel; Ribosomal RNA; Role; Sampling; Sequence Analysis; Site; Surveys; System; Temperature; Time; Universities; Vagina; Variant; Work; base; cost; design; flexibility; gene cloning; genome sequencing; improved; interest; metabolomics; microbial; preference; public health relevance; rRNA Genes

DESCRIPTION (provided by applicant): Analysis of ribosomal RNA (rRNA) genes is one of the most important methods for characterizing the organisms present in an environment, including those associated with medical conditions. The first step of these analyses is usually to apply the polymerase chain reaction (PCR) to specifically amplify the genes of interest. This requires the use of primers that are specific to the rRNA genes, yet capable of accommodating all of the natural variation of the genes. The investigators will analyze sequence variation in rRNA gene primer-binding sites in the currently available data with the goal of improving current primer designs to recognize the full range of relevant organisms. They will also explore trade-offs in using sequence degeneracy, noncanonical nucleotides, and primer annealing stringency to accommodate the observed sequence diversity. In so doing, they will explore the usefulness of strategies that might guide amplifications toward a common template and primer sequence. These goals will use computer analysis of sequences from the databases and experimental comparisons of alternative primer designs and amplification protocols. The results of much of the proposed work will be applicable for molecular analyses based on molecules other than rRNA genes. PUBLIC HEALTH RELEVANCE Microbes play important roles in human health and disease. There are increasing numbers of studies of the mixture of microbes associated with both normal and abnormal conditions. Most in-depth studies are based upon the analysis of particular molecules, most commonly the ribosomal RNAs. The goal of the proposed studies is to improve these methods in the detection of the full diversity of organisms present in the samples. This work will be integrated with on-going analyses of vaginal microbiota in humans.

描述（由申请人提供）：核糖体RNA（RRNA）基因的分析是表征环境中存在的生物（包括与医疗条件相关的生物）的最重要方法之一。这些分析的第一步通常是应用聚合酶链反应（PCR）以特异性扩增感兴趣的基因。这需要使用特定于rRNA基因的引物，但能够适应基因的所有自然变异。研究人员将分析当前可用数据中rRNA基因引物结合位点的序列变化，以改善当前底漆设计以识别各种相关生物。他们还将使用序列退化，非规范核苷酸和引物退火的严格性来探索权衡取舍，以适应观察到的序列多样性。这样一来，他们将探讨可能指导对通用模板和底漆序列的扩增的策略的实用性。这些目标将使用数据库中的序列分析以及替代引物设计和放大协议的实验比较。许多提出的工作的结果将适用于基于rRNA基因以外的分子的分子分析。公共卫生相关性微生物在人类健康和疾病中起重要作用。与正常情况和异常条件相关的微生物的混合物的研究越来越多。大多数深度研究基于对特定分子的分析，最常见的是核糖体RNA。拟议的研究的目的是改善这些方法在检测样品中存在的各种生物的多样性中。这项工作将与人类阴道微生物群的持续分析相结合。