IMPROVED METHODS FOR ISOLATING RNA FROM LIMITED SAMPLES

从有限样品中分离 RNA 的改进方法

• 批准号: 6212240
• 负责人: MARIANNA Mansfield GOLDRICK
• 金额: $ 22.09万
• 依托单位: AMBION, INC.
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-02-01 至 2002-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6212240
• 关键词: RNA; cytogenetics; expression cloning; flow cytometry; human tissue; lasers; method development; microarray technology; northern blottings; nucleic acid purification; nucleic acid quantitation /detection; tissue /cell culture

DESCRIPTION: A need currently exists for methods to quantitatively purify RNA from small-scale samples (1-10,000 cells and tissue amounts ranging from 0.05 - 5 mg). Recent technical advances in the ability to microdissect individual cell types by laser capture from thin tissue sections has led to a need for better methods to isolate RNA from a few cells for expression profiling. Also, there is increasing demand for RNA isolation from small tissue samples. Clinical specimens are becoming smaller, with a trend toward fine-needle aspirates replacing more invasive surgery. Another situation that may require RNA purification from limited samples is in assessing the cells sorted by FACS, such as in combinatorial screening strategies for identifying minor populations expressing desirable responses to pharmacological compounds. In all of these cases, isolation of the maximum possible yields of high quality intact RNA is a requirement and a challenge. Compounding the challenge is the difficulty in assessing RNA yields from limited samples; traditional methods based on spectrophotometric assessment and gel analysis are too insensitive. The goal of this application is to develop methods for efficient quantitative recovery of high-quality RNA from limited samples, especially from those obtained by laser capture microdissection. PROPOSED COMMERCIAL APPLICATION: The proposed project will result in kits for the research and clinical markets to use for isolation of RNA from micro-scale samples. It is anticipated that the kits will be especially useful for expression profiling of cells recovered using Laser Capture Microdissection.

描述：目前需要定量纯化 RNA 的方法 来自小规模样本（1-10,000 个细胞和组织数量范围为 0.05 - 5 毫克）。显微解剖单个细胞能力的最新技术进展 通过激光从薄组织切片中捕获类型导致需要更好的 从一些细胞中分离 RNA 进行表达谱分析的方法。还有，还有 对从小组织样本中分离 RNA 的需求不断增加。临床 标本变得越来越小，细针抽吸成为趋势 取代更具侵入性的手术。另一种可能需要RNA的情况 从有限的样品中纯化是为了评估通过 FACS 分选的细胞， 例如用于识别少数群体的组合筛查策略 表达对药理学化合物的期望反应。在所有这些 在这种情况下，最大可能产量的高质量完整 RNA 的分离是一个 要求和挑战。使挑战更加复杂的是困难 评估有限样本的 RNA 产量；传统方法基于 分光光度评估和凝胶分析太不灵敏。目标是 该应用旨在开发有效定量回收的方法 从有限的样品中提取高质量的 RNA，尤其是通过激光获得的样品 捕获显微解剖。 拟议的商业应用： 拟议的项目将产生用于研究和临床市场的试剂盒 从微型样品中分离 RNA。预计该套件将特别 可用于使用激光捕获显微切割回收的细胞的表达谱。