Genomic siRNA Libraries as Tools for Pseudo-Genetics

基因组 siRNA 文库作为伪遗传学工具

• 批准号: 6690950
• 负责人: DAVID M BROWN
• 金额: $ 25.45万
• 依托单位: AMBION, INC.
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-09-30 至 2004-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6690950
• 关键词: DNA; RNA; biotechnology; complementary DNA; flow cytometry; genetic library; genetic promoter element; polymerase chain reaction; technology /technique development; transfection; transfection /expression vector

DESCRIPTION (provided by applicant): The proposed research will enable mammalian cells to be used in mutation/selection experiments to identify genes involved in cellular processes. The objective of the proposal is to develop procedures for preparing siRNA libraries using genomic DNA and cDNA samples as the source material for siRNA templates. Libraries of siRNA vectors could comprise 1,000,000,000 to 1,000,000,000,000 unique siRNA expression vectors, making it possible to cover entire mammalian genomes with overlapping siRNAs. Mammalian cell populations transfected or transduced with siRNA vector libraries will be placed under selection to create cell populations expressing a desirable phenotype. The siRNA vector(s) integrated in the genomes of the selected cells will be amplified, cloned, and sequenced to reveal the siRNA template sequence. The siRNA sequences will be used to identify the genes whose down-regulation create the phenotype that was selected. The siRNA vector libraries should enable mutation/selection experiments to be used to correlate gene function and identity in mammalian cells.

描述（由申请人提供）：拟议的研究将使哺乳动物细胞能够在突变/选择实验中使用，以鉴定涉及细胞过程的基因。该提案的目的是制定使用基因组DNA和cDNA样品作为siRNA模板的源材料制备siRNA文库的程序。 siRNA矢量的库可以包括1,000,000,000至1,000,000,000,000千万siRNA表达矢量，从而可以用重叠的siRNA覆盖整个哺乳动物基因组。用siRNA矢量文库转染或转导的哺乳动物细胞群将被置于选择下，以创建表达理想表型的细胞群体。积分在选定细胞的基因组中的siRNA载体将被放大，克隆和测序以揭示siRNA模板序列。 siRNA序列将用于识别其下调产生所选表型的基因。 siRNA矢量文库应使突变/选择实验用于将基因功能和哺乳动物细胞的同一性相关联。