Equipment for Spatiotemporal Dynamics of the Genome by 3D Orbital Tracking

三维轨道跟踪基因组时空动态研究装置 项目摘要 我们的目标是开发3D轨道跟踪，一个强大的单分子荧光波动 技术，以测量基因激活的时间，剪接，和基因组组织在一个活跃的基因， 活细胞。通过在共聚焦显微镜中扫描荧光标记的转录位点周围的环形激光， 在显微镜下，我们可以在很长一段时间内高速测量活细胞的荧光强度。通过 荧光标记，DNA，RNA，和蛋白质因子在不同的颜色，我们可以监测复杂的组装， RNA的转录、剪接和终止，以及远端DNA元件如增强子的邻近， 活基因组 以新的方式或新的工具观察自然，往往会揭示出以前未知的细节。这项建议 描述了这样一个项目。尽管人们已经做了很多工作来研究转录、剪接和终止 他们还没有能够描述RNA是如何从 一个基因然后被释放或者当剪接发生时。有了这样一个工具，我们可以看到许多以前 隐形的我们可以开始辨别复杂的生化反应，这些反应只能在活细胞中测量。 该研究项目将支持跨学科的本科生，研究生和博士后学员，他们将 学习基因组生物学和单分子成像方面的先进技术， 转录和剪接的时空动力学。项目成员将参加各种活动， 在科学、数学、工程和技术方面代表性不足的群体的参与。

Equipment for Spatiotemporal Dynamics of the Genome by 3D Orbital Tracking Project Summary Our goal is to develop 3D orbital tracking, a powerful single-molecule fluorescence fluctuation technique, to measure the timing of gene activation, splicing, and genome organization at an active gene in living cells. By scanning a laser in a circle around a fluorescently labeled site of transcription in a confocal microscope, we can measure fluorescence intensity at high speed for a very long time in living cells. By fluorescent labeling, DNA, RNA, and protein factors in different colors, we can monitor complex assembly, transcription, splicing, and termination of RNA and the proximity of distal DNA elements like enhancers in the living genome. Looking at nature in a new way, or with a novel tool, often reveals previously unknown details. This proposal describes such a project. Although much work has been done to study transcription, splicing, and termination of RNA, they have yet to be able to characterize a complete kinetic profile of how the RNA is synthesized from a gene and then released or when splicing occurs. With such a tool, we may see much that was previously invisible. We can begin to discern complex biochemical reactions that can only be measured in living cells. This research project will support interdisciplinary undergraduate, graduate, and postdoctoral trainees, who will learn advanced techniques in genome biology and single-molecule imaging, shedding new light on the spatiotemporal dynamics of transcription and splicing. Project members will participate in activities to broaden the participation of underrepresented groups in science, mathematics, engineering, and technology.