Structure of genomic DNA at single-nucleotide resolution

描述（由申请人提供）：最近完成的人类基因组为深入了解人类有机体的功能提供了原始数据。对基因组序列的了解将带来治疗疾病和确保健康的新方法。然而，要理解遗传信息是如何被检索和表达的，需要的不仅仅是核苷酸序列。DNA的三维结构对于包装基因组和调节基因表达的蛋白质的功能至关重要，但目前还没有能够在基因组尺度上绘制DNA结构的方法。在此应用中，提出了一种新的测定基因组DNA结构信息的实验技术。这种结构信息将为DNA元素百科全书（ENCODE）项目产生的数据增加一个新的维度。本应用程序中概述的实验方法针对超过90%的不编码蛋白质的人类基因组，其中存在基因调控所需的信息。本文提出的绘制基因组DNA结构图的方法利用了DNA结构的高分辨率化学探针——羟基自由基。本应用程序的具体目的是：(1)设计和实现一个羟基自由基DNA切割模式数据库。高通量方法将用于收集大量的实验羟基自由基DNA切割数据。这些实验将使用DNA文库和ENCODE区DNA序列。(2)建立预测任何DNA序列羟基自由基切割模式的方法。羟基自由基切割模式数据库将用于构建DNA结构如何依赖于核苷酸序列的模型。该模型将用于预测基因组DNA的羟基自由基切割模式，包括部分人类基因组的ENCODE区域。(3)绘制基因组DNA结构图。由于先前的工作已经证明羟基自由基切割模式提供了DNA表面形状的详细信息，实验（和预测）羟基自由基切割模式将用于绘制对染色质折叠至关重要的顺式作用因子结合位点和序列的图谱。计算工具，包括隐马尔可夫模型，将在羟基自由基切割模式数据库上进行训练，以识别基因组DNA序列的结构模式。

DESCRIPTION (provided by applicant): The recent completion of the human genome has provided the raw data on which to build a deep understanding of how the human organism functions. New ways to attack diseases and ensure health will result from knowledge of genome sequences. However, more than just the sequence of nucleotides is needed to understand how genetic information is retrieved and expressed. The three-dimensional structure of DNA is critical to the functions of the proteins that package the genome and regulate gene expression, yet no method exists that is capable of mapping DNA structure on a genomic scale. In this application, a new experimental technology is proposed for determining structural information for genomic DNA. This structural information will add a new dimension to the data that will be produced by the Encyclopedia of DNA Elements (ENCODE) Project. The experimental approach outlined in this application is aimed at the more than 90% of the human genome that does not code for proteins, where the information that is necessary for gene regulation resides. The method proposed here for making maps of genomic DNA structure makes use of a high-resolution chemical probe of DNA structure, the hydroxyl radical. The Specific Aims of this application are: (1) To design and implement a database of hydroxyl radical DNA cleavage patterns. High throughput methods will be used to collect large amounts of experimental hydroxyl radical DNA cleavage data. DNA libraries and ENCODE region DNA sequences will be used for these experiments. (2) To develop methods to predict the hydroxyl radical cleavage pattern of any DNA sequence. The hydroxyl radical cleavage pattern database will be used to construct a model for how the structure of DNA depends on the sequence of nucleotides. This model will be used to predict the hydroxyl radical cleavage patterns of genomic DNA, including parts of the ENCODE regions of the human genome. (3) To make structural maps of genomic DNA. Since previous work has demonstrated that the hydroxyl radical cleavage pattern provides detailed information on the shape of the surface of DNA, experimental (and predicted) hydroxyl radical cleavage patterns will be used to make maps of cis-acting factor binding sites and sequences critical to the folding of chromatin. Computational tools, including hidden Markov models, will be trained on the database of hydroxyl radical cleavage patterns in order to recognize structural patterns in genomic DNA sequences.