Flow Cytometry: Digital Processing of Molecular Information

流式细胞术：分子信息的数字处理

• 批准号: 9513602
• 负责人: David Galbraith
• 金额: $ 5万
• 依托单位: University of Arizona
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 1996
• 资助国家: 美国
• 起止时间: 1996-03-01 至 1997-02-28
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=9513602&HistoricalAwards=false
• 关键词: Flow; Cytometry; Digital; Processing; Molecular

This project involves the design, implementation, and evaluation of flow cytometric instrumentation and operating procedures for use in the cytoenzymological and histochemical analysis of eukaryotic cells. It is divided into two subprojects. The first involves the development of a second-generation digital data acquisition and pulse processing system for the real-time analysis and sorting of cells based on pulse shape features. The second subproject involves the development of novel gene expression systems for use in the molecular and cellular analysis in flow of transgenic eukaryotic cells. We then intend to employ these techniques for the selection through flow sorting of transformed plant nuclei according to the level of transgene expression, ultimately directed at the construction of cDNA libraries representing the developmentally-regulated genes of tissues that are microscopically-small. The significance of this work is as follows. In the first case, our previous work has shown that important information about eukaryotic cells is contained within the waveform shapes produced by these cells produced when they are subjected to analysis using flow cytometry. Present flow cytometric instrumentation cannot extract this shape-based information. The capability of the proposed second generation machine to analyze and sort cells based on pulse shape information will provide important knowledge about cellular substructure. This will lead to improved understanding of the basic cellular biology of living cells. In the second case, the ability of scientists to analyze the expression of genes within different cell types is hampered when these cells are interspersed within other cell types. The molecular methods proposed in this project should provide general means for isolation of transcripts from any desired cell type. This should greatly facilitate advances in the study of developmental- and cell type-specific gene expression.

该项目涉及流式细胞仪仪和操作程序的设计，实施和评估，以用于真核细胞的细胞合理学和组织化学分析。 它分为两个子标记。第一个涉及开发第二代数字数据采集和脉冲处理系统，以基于脉冲形状特征对单元进行实时分析和分类。第二次次要介绍涉及开发新型基因表达系统，用于在转基因真核细胞流中用于分子和细胞分析。然后，我们打算采用这些技术来选择转化的植物核根据转基因表达的水平进行选择，最终是针对代表微小组织组织的发育中调节基因的cDNA文库的构建。 这项工作的意义如下。在第一种情况下，我们以前的工作表明，有关真核细胞的重要信息包含在使用流式细胞仪进行分析时产生的这些细胞产生的波形形状中。 当前的流式细胞仪仪无法提取此基于形状的信息。拟议的第二代机器根据脉冲形状信息进行分析和分类细胞的能力将提供有关细胞子结构的重要知识。这将导致人们对活细胞的基本细胞生物学的理解。 在第二种情况下，当这些细胞插入其他细胞类型中时，科学家分析不同细胞类型中基因表达的能力会受到阻碍。该项目中提出的分子方法应提供从任何所需细胞类型分离转录本的一般手段。这应该极大地促进发展性和细胞类型特异性基因表达的进展。