Accessible high-throughput single-cell genome sequencing

可获得的高通量单细胞基因组测序

基本信息

批准号：
10612457
负责人：
Andrew Adey
金额：
$ 37.3万
依托单位：
OREGON HEALTH & SCIENCE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-04-21 至 2025-03-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10612457
关键词：
ATAC-seq Acoustics Area Atlases Bar Codes Basic Science Benchmarking Biological Assay Cancer Biology Catalogs Cell Line Cell Nucleus Cells Chemistry Chromium Clinical Collaborations Communities Complex Computer software Cost Effectiveness Analysis Cost Savings Custom DNA Sequence Rearrangement Data Detection Evolution Genes Genetic Genome Genomics High-Throughput Nucleotide Sequencing Human Hybrids In Situ Libraries Liquid substance Metastatic breast cancer Methodology Methods Microfluidics Mus Nucleosomes Pancreatic Ductal Adenocarcinoma Point Mutation Population Primary Brain Neoplasms Property Protocols documentation Publishing Reagent Reproducibility Research Resistance Sampling Single Nucleotide Polymorphism Solid Neoplasm Structure Technology Therapeutic Validation analysis pipeline anticancer research cost cost effective density design exome exome sequencing experimental study gene panel genome sequencing genomic profiles improved in situ sequencing indexing instrument instrumentation interest nano novel single cell sequencing therapy resistant tool tumor tumor initiation tumor progression variant detection whole genome

项目摘要

PROJECT SUMMARY Initiation and evolution of most solid tumors involve complex genomic rearrangements in addition to point mutations – all of which can contribute to cancer progression and to the evolution of resistance to therapeutics. Single-cell whole genome sequencing (scWGS) has proven invaluable for the identification of tumor subpopulations and in advancing our understanding of clonal dynamics and tumor evolution. Despite the value of scWGS, there is a dearth of commercially-available options that provide enough cell throughput (i.e. power) and/or genomic coverage to fully catalogue and characterize clonal populations within a tumor sample. This can be particularly problematic when rare, possible therapy-resistant, subpopulations are present within a tumor that may elude detection using existing methodologies. Here we will solve the accessibility problem by developing technologies that leverage widely-available instrumentation and off-the-shelf reagents. We previously described proof-of-principle technologies to construct scWGS libraries contained within the nucleus (in situ) of each cell that are then carried through cell barcoding using our custom workflow. We will extend, develop, and adapt these technologies to provide three workflows that will meet the needs of the cancer research community by providing readily-accessible order-of-magnitude improvements to cell throughput, cell coverage and cost savings for scWGS analysis over current options. The first is focused on high cell throughput using widely available instrumentation; the second leverages our high- content chemistry to produce high-coverage single cell genomic profiles with reduced reagents and improved throughput; and finally, the third approach implements target capture to target exonic sequence or other regions of interest to reduce sequencing costs. Each of these is targeted to meet the needs of specific areas of cancer research, from basic science to clinical, and share the theme of accessibility.

项目摘要大多数实体瘤的起始和进化涉及复杂的基因组重排，除了点突变 - 所有这些都可以导致癌症进展和抗治疗的耐药性演变。单细胞全基因组测序（SCWG）已被证明是鉴定肿瘤的宝贵的宝贵亚种群并促进我们对克隆动力学和肿瘤进化的理解。尽管有价值在SCWGS中，有商业上可用的选项死亡，可提供足够的细胞吞吐量（即功率）和/或基因组覆盖范围，以完全分类并表征肿瘤样品中克隆种群。这当罕见的，可能抗治疗的亚群时，可能会特别有问题可能使用现有方法洗脱检测的肿瘤。在这里，我们将通过开发利用广泛可用的技术来解决可访问性问题仪器和现成的试剂。我们先前描述了原理验证技术以构建 SCWGS文库包含在每个细胞的细胞核（原位）中，然后通过细胞条形码携带使用我们的自定义工作流程。我们将扩展，开发和适应这些技术以提供三个工作流程这将通过提供易于接近的速度命令来满足癌症研究界的需求改善细胞吞吐量，细胞覆盖范围和SCWGS分析的成本节省，而不是当前选项。这首先使用广泛可用的仪器专注于高细胞吞吐量；第二个利用了我们的高含量化学以产生具有减少试剂并改进的高覆盖范围的单细胞基因组谱吞吐量；最后，第三种方法将目标捕获实现到目标外显子序或其他降低测序成本的兴趣区域。这些都针对的是满足特定领域的需求癌症研究，从基础科学到临床，并具有可访问性的主题。