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.

该项目涉及用于真核细胞的细胞酶学和组织化学分析的流式细胞仪仪器和操作程序的设计、实施和评价。 它分为两个分项目。第一个项目涉及开发第二代数字数据采集和脉冲处理系统，用于根据脉冲形状特征对细胞进行实时分析和分选。第二个子项目涉及开发新的基因表达系统，用于转基因真核细胞流中的分子和细胞分析。然后，我们打算采用这些技术，根据转基因表达水平，通过流式分选转化的植物细胞核进行选择，最终针对代表显微镜下小的组织的发育调控基因的cDNA文库的构建。 这项工作的意义如下。在第一种情况下，我们以前的工作已经表明，有关真核细胞的重要信息包含在这些细胞产生的波形形状中，当它们进行流式细胞术分析时。 目前的流式细胞仪无法提取这种基于形状的信息。所提出的第二代机器的能力，分析和分类细胞的脉冲形状信息的基础上，将提供重要的知识细胞的子结构。这将导致对活细胞的基本细胞生物学的更好理解。 在第二种情况下，当这些细胞散布在其他细胞类型中时，科学家分析不同细胞类型中基因表达的能力受到阻碍。在这个项目中提出的分子方法应提供一般手段，从任何所需的细胞类型的转录分离。这将极大地促进发育和细胞类型特异性基因表达研究的进展。