In vivo transcription factor DNA binding analysis

体内转录因子 DNA 结合分析

• 批准号: 6643757
• 负责人: JOHN ARCHDEACON
• 金额: $ 9.99万
• 依托单位: ACTIVE MOTIF, INC.
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-04-01 至 2003-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6643757
• 关键词: DNA; chromatin; immunoprecipitation; intermolecular interaction; microarray technology; nuclear factor kappa beta; protein quantitation /detection; reagent /indicator; technology /technique development; transcription factor

DESCRIPTION (provided by applicant): Currently, there exist a number of methods for the study of transcription factors and gene-promoter activity. These include reporter gene assays, gel shift assays, and expression-profiling methods such as microarray analysis and SAGE. These methods are well established and have been used to generate a wealth of information. Accordingly, there are many commercially available kits designed to facilitate and speed these analyses. However, none of the above methods enables identification of the DNA targets that are actually bound by a particular transcription factor in a particular population of cells. The current best method to generate this type of transcription "snapshot" is the Chromatin Immunoprecipitation (CHIP) assay. This method is technically demanding and is not well supported by commercially available complete kits. Similarly, a powerful extension of the CHIP assay, specifically, the merging of the CHIP assay with microarray analysis of the immunoprecipitated DNA, is not available commercially. We propose to develop reagents to simplify and extend use of the CHIP assay for the study of transcription factor/DNA interactions in living mammalian cells. The long-term objective of this proposal is the development and commercialization of novel kits and reagents that speed the study of in vivo interactions between transcription factors and DNA.

描述（申请人提供）：目前，转录因子和基因启动子活性的研究方法有很多。这些方法包括报告基因分析、凝胶移位分析和表达谱分析方法，如微阵列分析和SAGE。这些方法得到了很好的确立，并已被用来产生丰富的信息。因此，有许多商业上可用的试剂盒，旨在促进和加快这些分析。然而，上述方法都无法识别特定细胞群中实际由特定转录因子结合的DNA靶标。目前生成这种类型的转录“快照”的最佳方法是染色质免疫沉淀（CHIP）测定。这种方法在技术上要求很高，而且商业上可用的完整工具包不支持这种方法。同样，CHIP测定法的强大扩展，特别是CHIP测定法与免疫沉淀DNA的微阵列分析的合并，在商业上是不可用的。我们建议开发试剂来简化和扩展CHIP测定在活体哺乳动物细胞中转录因子/DNA相互作用研究的使用。该提案的长期目标是开发和商业化新的试剂盒和试剂，以加快转录因子和DNA之间体内相互作用的研究。