Nanopore based profiling of epigenetic state

项目摘要： 尽管参考人类基因组和甚至表观基因组的极好表征（ENCODE，Epigenetics 路线图，4D Nucleome项目）已经产生，我们仍然缺乏一个清晰的认识，如何不同的 表观基因组的各个方面共同控制基因的表达。随着经济实惠的DNA测序技术的出现 技术，方法已经开发用于检查核组织，蛋白质-DNA相互作用， 染色质可及性和甲基化状态。但这些方法通常只询问一个方面， 表观基因组的读取通常太短而不能提供关键的相关信息。我们建议 开发一种新的表观遗传表征方法，在这一轮的资金集中在蛋白质- 通过利用纳米孔测序可检测的长读段和修饰碱基的DNA相互作用 平台我们将增强用于分析蛋白质-DNA相互作用的细微修饰的信号。 我们正在开发一种在蛋白质-DNA结合位点附近插入序列标签（nanoTUBI）的方法。我们 我将实施这些方法，重点是长读序测序，深入了解长相关性 测量和迄今未探索的重复区域，定相整个表观基因组。这些工具和 分析管道将为基因调控机制提供新的见解， 人类疾病

Project Summary: Though a reference human genome and even excellent characterization of the epigenome (ENCODE, Epigenetics Roadmap, 4D Nucleome Project) has been generated, we still lack a clear understanding of how the different facets of the epigenome collaborate to control gene expression. With the advent of affordable DNA-sequencing technologies, methods have been developed for examining nuclear organization, protein-DNA interaction, chromatin accessibility, and methylation state. But these methods generally interrogate only one aspect of the epigenome at a time and reads are typically too short to provide critical correlative information. We propose the development of a novel epigenetic characterization methodology, in this round of funding focusing on protein- DNA interaction through leveraging the long reads and modified bases detectable with a nanopore sequencing platform. We will enhance the signal from subtle modifications being used to profile protein-DNA interactions. We are developing a method to insert sequence tags (nanoTUBI) near the sites of protein-DNA binding. And we will implement these methods focusing on long read sequencing, gaining insights into long correlative measurements and heretofore unexplored repetitive areas, phasing the entire epigenome. These tools and analysis pipelines will grant new insights into mechanisms of gene regulation, as well as their implications for human disease.