A next-generation method for cytogenomics using Hi-C proximity ligation sequencing

使用 Hi-C 邻近连接测序的下一代细胞基因组学方法

• 批准号: 10397703
• 负责人: Stephen Matthew Eacker
• 金额: $ 83.79万
• 依托单位: PHASE GENOMICS, INC.
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-05-01 至 2023-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10397703
• 关键词: Accounting; Address; Affect; Aneuploidy; Animals; Benchmarking; Cell Line; Child; Chromosome abnormality; Chromosomes; Clinical; Collection; Computer software; Conceptions; Couples; Cytogenetic Analysis; Cytogenetics; DNA Ligation; DNA Sequence Rearrangement; DNA sequencing; Data; Data Set; Decision Making; Detection; Diagnosis; Diagnostic; Diagnostic Procedure; Event; Fetal Viability; Fluorescent in Situ Hybridization; Funding; Future; Gametogenesis; Genetic; Genome; Genomics; Goals; Grant; Hi-C; Industry Standard; Infertility; Karyotype; Karyotype determination procedure; Laboratories; Lead; Length; Lesion; Letters; Ligation; Manuals; Methodology; Methods; Molecular Biology; Mosaicism; Mutation; Online Systems; Outcome; Performance; Phase; Plants; Pregnancy loss; Process; Protocols documentation; Reagent; Reporting; Reproducibility; Reproduction; Reproductive Medicine; Research; Resolution; Sampling; Sensitivity and Specificity; Series; Services; Single Nucleotide Polymorphism; Small Business Innovation Research Grant; Software Tools; Study Section; System; Technology; Testing; Time; United States; Variant; clinically relevant; cloud based; cohort; computational platform; cost; design; feasibility testing; genetic testing; improved; large datasets; meetings; next generation; novel strategies; precision medicine; reproductive; scaffold; tool

ABSTRACT Reproductive genetic tests are a disparate collection of methodologies with identify specific classes of mutations known to interfere with gametogenesis, conception and fetal viability. The number and variety of tests addressing this clinical need reflect the scale of genetic lesions that are relevant to reproductive medicine, ranging from whole chromosome aneuploidy to single nucleotide variants. In this proposal, we describe a highly scalable reproductive genetic test that can deliver results that span this scale at high resolution, lower cost, and faster turnaround time than current tests. Our approach uses a proximity ligation DNA sequencing method called Hi-C, which captures ultra-long-range genomic contiguity information using ubiquitious short-read sequencing and benchtop molecular biology. We have demonstrated extensively that Hi-C data can be used to order and orient genome assemblies, reconstructing end-to-end chromosome sequences. This method also identifies structural genomic rearrangements including balanced translocations, inversions, and other aberrations not detectable by typical sequencing approaches. We propose to apply high-throughput proximity ligation as a cytogenomic method to detect the breadth of chromosomal aberrations at high resolution and low cost. This proposal outlines a path to a commercially available product and service, which will establish a highly validated method for use in research and eventually in a diagnostic setting. This will be accomplished by 1) designing an easy to use Hi-C protocol amenable to multiwell plate handling, 2) building a robust computational platform to reproducibly call chromosome aberrations from Hi-C data, and 3) proving the validity and reproducibility of these methods on real world samples. The resulting kit and software product will be a new cytogenomic method called Karyotyping by SequencingTM (KBS) that we will deploy as a Precision Medicine test for the reproductive testing market.

抽象的 生殖遗传测试是具有识别特定类别突变类别的方法集合 已知会干扰配子发生，受孕和胎儿生存能力。解决的测试数量和多样性 这种临床需求反映了与生殖医学相关的遗传病变的规模，从 整个染色体非整倍性到单核苷酸变体。在此提案中，我们描述了一个高度可扩展的 生殖基因测试可以提供以高分辨率，较低成本和更快的范围跨越该量表的结果 周转时间比当前测试。 我们的方法使用称为HI-C的接近连接DNA测序方法，该方法捕获了超长范围 基因组连续性信息，使用无处不在的短阅读测序和台式分子生物学。我们 已经广泛证明了HI-C数据可用于订购和定向基因组组件， 重建端到端染色体序列。该方法还鉴定了结构基因组 重排，包括平衡易位，反转和其他畸变，无法通过典型检测到 测序方法。 我们建议将高通量接近连接作为一种细胞基因组方法来检测广度 高分辨率和低成本的染色体畸变。该提议概述了商业上的途径 可用的产品和服务将建立一种高度验证的研究方法，并最终 在诊断环境中。这将通过1）设计易于使用的HI-C协议来完成 多井板处理，2）构建一个可重复调用染色体畸变的强大计算平台 从HI-C数据和3）证明这些方法对现实世界样本的有效性和可重复性。这 由此产生的套件和软件产品将是一种新的细胞基因组方法，称为序列化的核分型（KBS）（KBS） 我们将作为生殖测试市场的精确医学测试部署。