Rapid Quantitation of Small RNA

小 RNA 的快速定量

• 批准号: 7231335
• 负责人: GARY J LATHAM
• 金额: $ 44.1万
• 依托单位: AMBION DIAGNOSTICS, INC.
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-08-15 至 2009-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7231335
• 关键词: Adopted; Base Sequence; Basic Science; Biological; Biological Assay; Class; Clinical; Complementary DNA; Condition; Cultured Cells; Data; Detection; Development; Diagnostic; Diagnostics Research; Discrimination; Disease Progression; Endoribonucleases; Fresh Tissue; Gene Expression; Genes; Goals; Health; Heart; Human; Individual; Institution; Liquid substance; Malignant Neoplasms; Measures; Methods; MicroRNAs; Molecular Profiling; Northern Blotting; Pancreatic ribonuclease; Phase; Play; Polymerase; Polymerase Chain Reaction; Procedures; Publications; RNA; Reaction; Reagent; Research; Research Personnel; Reverse Transcriptase Polymerase Chain Reaction; Ribonucleases; Role; Sampling; Sensitivity and Specificity; Slice; Small RNA; System; Techniques; Testing; Therapeutic; Therapeutic Intervention; Tissues; Validation; Virus Diseases; Work; base; calin; design; improved; multiplex detection; nervous system disorder; prognostic; tool

DESCRIPTION (provided by applicant): MicroRNAs (miRNAs) are a significant new class of biomolecules with potential to enhance human health. In the last three years, miRNA publications have increased exponentially as researchers have discovered the potential importance of these small RNAs in human cancer, neurological disease, and viral infection. Ambion researchers have worked to develop the tools necessary to understand the relationship between miRNAs and human health. Our results and the results from other institutions indicate that miRNAs may provide useful markers for the development of diagnostic and prognostic assays. In addition, there are hints that miRNAs might play an active role in disease progression, making them candidates for therapeutic intervention. The goal of this proposal is to develop rapid, sensitive, high-throughput methods to quantify the levels of small RNA molecules such as microRNAs (miRNA) in biological samples. These small RNAs have emerged as important regulators of gene expression which are critical for development and disease progression. Unfortunately, there currently are no rapid, quantitative methods available to measure levels of miRNA in clinical samples. The proposed research will result in the development of kits and reagents to rapidly quantify miRNAs in samples derived from tissues or cell culture. During Phase I, we invented a qRT-PCR based method to convert target miRNA molecules into cDNA and which are then amplified and detected using PCR. This method enables the specific detection of individual miRNAs in RNA samples. In Phase II we propose to optimize the method to facilitate its commercial use in research and diagnostic applications. Our Phase II research will improve sensitivity and specificity, permit simultaneous detection of multiple miRNAs in a single reaction, and enable detection on a Luminex liquid bead array platform. These improvements will enable rapid miRNA profiling for basic research, clinical validation of diagnostic and therapeutic miRNA candidates, and put a system in place for rapid development of miRNA-based clinical diagnostic assays.

描述（由申请人提供）：MicroRNA (miRNA) 是一类重要的新型生物分子，具有增强人类健康的潜力。在过去三年中，随着研究人员发现这些小 RNA 在人类癌症、神经系统疾病和病毒感染中的潜在重要性，miRNA 的出版物呈指数级增长。 Ambion 研究人员致力于开发了解 miRNA 与人类健康之间关系所需的工具。我们的结果和其他机构的结果表明 miRNA 可能为诊断和预后分析的开发提供有用的标记。此外，有迹象表明 miRNA 可能在疾病进展中发挥积极作用，使其成为治疗干预的候选者。该提案的目标是开发快速、灵敏、高通量的方法来量化生物样品中小 RNA 分子（例如 microRNA (miRNA)）的水平。这些小RNA已成为基因表达的重要调节因子，对发育和疾病进展至关重要。不幸的是，目前尚无快速、定量的方法可用于测量临床样本中 miRNA 的水平。拟议的研究将开发出试剂盒和试剂，以快速定量组织或细胞培养物样本中的 miRNA。在第一阶段，我们发明了一种基于 qRT-PCR 的方法，将目标 miRNA 分子转化为 cDNA，然后使用 PCR 进行扩增和检测。该方法能够特异性检测 RNA 样品中的单个 miRNA。在第二阶段，我们建议优化该方法，以促进其在研究和诊断应用中的商业用途。我们的 II 期研究将提高灵敏度和特异性，允许在单个反应中同时检测多个 miRNA，并能够在 Luminex 液珠阵列平台上进行检测。这些改进将使快速 miRNA 分析能够用于基础研究、诊断和治疗 miRNA 候选物的临床验证，并为快速开发基于 miRNA 的临床诊断分析建立一个系统。