Use of Light to Footprint DNA in Vitro and in Vivo

使用光在体外和体内足迹 DNA

• 批准号: 8805094
• 负责人: Michael Becker
• 金额: $ 29.8万
• 依托单位: University of Pittsburgh
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 1988
• 资助国家: 美国
• 起止时间: 1988-08-01 至 1992-01-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=8805094&HistoricalAwards=false
• 关键词: Use; Light; Footprint; DNA; Vitro

The ultimate goal of this research is to visualize, in intact cells, the structure of any gene. When gene expression is regulated at the level of transcription, changes in the structure of the gene are responsible for inducing changes in gene expression. These functional differences in gene structure are initiated and maintained by the interaction of regulatory proteins and chromosomal proteins with the gene, and possibly, by the formation of altered structures within the DNA helix. Dr. Becker is developing a set of photochemical methods, which, when taken together, is allowing one to detect as well as specify those elements of a gene's structure that are responsible for regulating its activity. This approach is based on earlier work which demonstrated that ultraviolet light can detect, at single nucleotide resolution, protein DNA contacts in whole cells. By directly probing gene structure in vivo rather than in vitro, potentially inaccurate or incomplete conclusions regarding the role of gene structure in regulating gene expression will be avoided. The ability to rapidly sequence any gene has revolutionized biology. Here the next level of organization, the structure of the gene, is being attacked. By determining the protein - DNA interactions and altered DNA structures within a gene general organizational principles for rendering the genetic message biologically active, stable, and inheritable will be uncovered.

这项研究的最终目标是在完整的细胞中可视化任何基因的结构。当基因表达在转录水平上被调节时，基因结构的变化就是导致基因表达变化的原因。基因结构中的这些功能差异是由调节蛋白和染色体蛋白与基因的相互作用启动和维持的，也可能是通过在DNA螺旋内形成改变的结构来启动和维持的。贝克尔博士正在开发一套光化学方法，当这些方法结合在一起时，人们可以检测并确定基因结构中负责调节其活动的那些元素。这种方法是基于早期的工作，证明了紫外线可以在单核苷酸分辨率下检测到整个细胞中的蛋白质DNA接触。通过直接探测体内而不是体外的基因结构，将避免关于基因结构在调节基因表达中的作用的潜在不准确或不完整的结论。对任何基因进行快速测序的能力使生物学发生了革命性的变化。在这里，下一个层次的组织，即基因的结构，正在受到攻击。通过确定蛋白质-DNA相互作用和基因内改变的DNA结构，将揭示使遗传信息具有生物活性、稳定性和可遗传性的一般组织原则。