Transcription indication reporter array technology

转录指示报告基因阵列技术

• 批准号: 6739251
• 负责人: XIN JIANG
• 金额: $ 17.44万
• 依托单位: PANOMICS, INC.
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-01-01 至 2004-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6739251
• 关键词: biotechnology; genetic library; genetic regulatory element; genetic transcription; high throughput technology; microarray technology; nucleic acid hybridization; nucleic acid probes; nucleic acid purification; nucleic acid sequence; reporter genes; technology /technique development; transcription factor; transfection

DESCRIPTION (provided by applicant): Transcription factors (TFs) are cellular proteins responsible for controlling gene expression. Understanding the mechanism underlying the activation of these proteins will help us dissect cellular signaling pathways and determine the relationship between transcription activation and human diseases. Monitoring the activities of transcription factors in vivo usually relies on a genetic reporter construct that contains a cis-element corresponding to a specific transcription factor, a minimal promoter, and a reporter gene. When the transcription factor is activated, it binds to the cis-element and induces the expression of the reporter gene. Through enzymatic analysis of the reporter, the activity of the transcription factor can therefore be measured, and furthermore the level of transcriptional induction can be determined by comparison of control and treated cells. As the reporter is a single molecule, each assay can determine only one transcription factor's activity. Consequently, if multiple transcription factors need to be measured, the work will be tedious and time-consuming because it will involve multiple transfections and a repeat of the same assay over and over again. In the study, we propose to use a series of tag sequences as the reporter. We will create a library of reporter constructs, each containing a cis-element and a unique genetic tag sequence. The expression of reporter tags will be induced if their corresponding transcription factors are activated and bound to the cis-elements in host cells. The expressed tag sequences can then be determined with an array assay. With this technology, the simultaneous activation of multiple transcription factors in signal transduction pathways in vivo can be monitored at once.

描述(由申请人提供)： 转录因子(TF)是负责控制基因表达的细胞蛋白质。了解这些蛋白激活的机制将有助于我们剖析细胞信号通路，并确定转录激活与人类疾病之间的关系。监测体内转录因子的活性通常依赖于包含与特定转录因子对应的顺式元件、最小启动子和报告基因的基因报告结构。当转录因子被激活时，它与顺式元件结合，诱导报告基因的表达。通过对报告分子的酶分析，可以测量转录因子的活性，进而可以通过对照细胞和处理细胞的比较来确定转录诱导的水平。由于该报告是一个单分子，每次检测只能确定一个转录因子的活性。因此，如果需要测量多个转录因子，这项工作将是乏味和耗时的，因为它将涉及多个转基因和反复重复相同的检测。在这项研究中，我们建议使用一系列标签序列作为报告。我们将创建一个报告结构文库，每个文库包含一个顺式元件和一个唯一的遗传标签序列。如果相应的转录因子被激活并与宿主细胞中的顺式元件结合，就会诱导报告标签的表达。表达的标签序列然后可以用阵列分析来确定。利用这项技术，可以同时监测体内信号转导通路中多个转录因子的同时激活情况。