High-content single-cell epigenetic technologies scalable to the human brain

高内涵单细胞表观遗传技术可扩展到人脑

• 批准号: 10369335
• 负责人: Andrew Adey
• 金额: $ 193.9万
• 依托单位: OREGON HEALTH & SCIENCE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-17 至 2024-09-16
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10369335
• 关键词: ATAC-seq; Atlases; Autopsy; BRAIN initiative; Bar Codes; Biological Assay; Brain; Catalogs; Cell Count; Cells; Cellular Assay; Censuses; Chemistry; Chromatin; Complement; DNA; DNA Methylation; Data; Data Set; Development; Epigenetic Process; Genetic Transcription; Goals; Hippocampus (Brain); Histones; Human; Hybrids; Individual; Libraries; Maps; Methods; Methylation; Molecular; Molecular Conformation; Molecular Profiling; Mus; Nucleic Acid Regulatory Sequences; Ploidies; Preparation; Production; Property; RNA; Regulatory Element; Sampling; Techniques; Technology; Time; Transposase; Variant; Visual Cortex; XCL1 gene; area striata; base; brain tissue; cell type; combinatorial; cost; experimental study; genome sequencing; genome-wide; histone modification; improved; indexing; interest; multiple omics; novel; single cell technology; technology development; tool; whole genome

PROJECT ABSTRACT To meet the goal of the BRAIN Initiative Cell Census Network to catalogue and produce molecular profiles of every cell type in the human brain; order-of-magnitude improvements in single-cell assay throughput and coverage are required. In this proposal, we detail strategies to meet these needs by leveraging novel chemistry workflows and combinatorial indexing techniques. This will include the development of two complementary techniques (s3 and s4) that overcome the major limitations of current technologies to produce far higher coverage of DNA-based properties, including chromatin accessibility (s3-ATAC) and histone marks (s4-CAT). These technologies will also be leveraged to develop assays that capture RNA transcription alongside the DNA-encoded property. We will also extend these workflows to produce order-of-magnitude improvements in cell coverage for our previously-described technology to profile single-cell DNA methylation in high throughput (sci-MET). The improved coverage will also enable the development of targeted capture for pooled single-cell DNA methylation libraries to reduce sequencing costs per cell while profiling regions of interest. These capture libraries will enable much larger cell datasets that can complement whole-genome single-cell profiles from a smaller sampling of cells. Finally, we will demonstrate these technologies by producing a preliminary atlas of epigenetic profiles in the human primary visual cortex and hippocampus to serve as a starting point for the large scale coordinated efforts of the BICCN.

项目摘要 为了实现BRAIN Initiative Cell Census Network的目标， 人类大脑中的每种细胞类型;单细胞测定通量的数量级改进， 覆盖是必需的。在这份提案中，我们详细介绍了通过利用新的化学方法来满足这些需求的策略。 工作流程和组合索引技术。这将包括发展两个互补的 技术（s3和s4）克服了现有技术的主要局限性， 覆盖基于DNA的特性，包括染色质可及性（s3-ATAC）和组蛋白标记（s4-CAT）。 这些技术也将被用来开发检测方法，捕获RNA转录，同时检测RNA的转录。 DNA编码的属性。我们还将扩展这些工作流程，以实现以下方面的数量级改进： 我们先前描述的以高通量分析单细胞DNA甲基化的技术的细胞覆盖率 （sci-MET）。改进的覆盖率还将使针对合并的单细胞的靶向捕获的发展成为可能。 DNA甲基化文库，以降低每个细胞的测序成本，同时分析感兴趣的区域。这些捕获 文库将使更大的细胞数据集，可以补充全基因组单细胞概况， 更小的细胞采样。最后，我们将通过制作一个初步的地图集来展示这些技术， 在人类初级视觉皮层和海马中的表观遗传谱作为研究的起点。 BICCN的大规模协调努力。