权益分类	功能权益	普通用户	{{item.name}}会员
{{category.name}}	{{benefitItem.name}}

Decoding chromosome structure with multiplexed super-resolution microscopy

用多重超分辨率显微镜解码染色体结构

基本信息

批准号：
9762943
负责人：
Peng Yin
金额：
$ 54.67万
依托单位：
HARVARD UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-09-01 至 2022-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9762943
关键词：
3-Dimensional Address Adopted Architecture Binding Biochemical Bioinformatics Biological Assay Biology Caliber Cell Nucleus Cells Chromatin Chromatin Fiber Chromatin Loop Chromosome Structures Chromosome Territory Chromosomes Color Crowding DNA DNA Damage DNA Probes DNA Repair DNA biosynthesis DNA-Protein Interaction Defect Detection Development Disease Environment Epigenetic Process Event Family Fluorescent in Situ Hybridization Gene Expression Profile Genome Genomic DNA Genomics Human Image Imagery Immunofluorescence Immunologic In Situ Individual Label Lead Length Malignant Neoplasms Maps Medical Methods Microscopy Modeling Nuclear Oligonucleotides Play Population Positioning Attribute Process Property Proteins RNA Research Research Personnel Resolution Role Sampling Site Specificity Structure Techniques Technology Thick Time Transcriptional Regulation Variant Visual Work X Inactivation base cell type chromosome conformation capture design developmental disease imaging modality improved insight meter molecular scale nanoscale programs single cell technology single molecule three dimensional structure tool

项目摘要

Summary Decades of study have revealed that genome organization is non-random and critically impacts many nuclear processes including the regulation of transcription, DNA replication, and DNA repair, and increasing evidence suggests that the three-dimensional structures adopted by chromosomes are critical for development and are often perturbed in disease. Much of our current understanding comes from biochemical techniques performed on large populations of cells, leading to many gaps in our understanding of the mechanisms that establish and maintain organizational states, particularly in the context of individual cells. We propose to introduce a new set of single-cell technologies based on the single-molecule super-resolution imaging method DNA-PAINT to bridge this gap with a suite of tools possessing both high multiplexibility and spatial resolution. Specifically, in Aim 1 we will develop a multiplexed (>20 color) super-resolution chromosomal imaging strategy to image genomic targets ranging from kilobases to multiple megabases in Iength, which will enable us to investigate the folding properties of the chromatin fiber in single cells over a range of length-scales. In Aim 2, we will develop multiplexed assays to co-localize proteins, RNA molecules, and specific genomic sites in individual cells at the nanoscale. We will then investigate organization of architectural proteins at a model hub of large chromatin loops on the human inactive X-chromosome. In Aim 3, we will develop a proximity-dependent super-resolution method to probe specific interactions between protein and DNA targets that will allow for the sensitive detection of molecular interactions in crowded environments. We will deploy this technology to query the composition and epigenetic states of the aforementioned chromatin looping hub in individual cells. Collectively, our methods will make many questions about the positioning, composition, and epigenetic states of specific genomic loci in individual cells accessible to researchers for the first time, and promise to impact diverse fields beyond chromosome biology.

总结