Massively Parallel Contiguity Mapping

描述（由申请人提供）：即使新技术继续降低DNA测序的成本，我们也迫切需要同样强大的方法来告知长距离邻接，以支持从头基因组组装和单倍型解析的基因组重测序。通过该计划的资助，我们探索了低成本、大规模并行捕获邻近信息的多种方法。我们的进展是实质性的，包括原位文库构建和光学测序的方法的开发，一种方法，其中我们利用“接触概率图”来产生第一个染色体规模的从头哺乳动物基因组组件完全基于短读段，和一种方法，该方法结合了连续性保持转座和组合索引，用于准确的，兆碱基规模的单倍型分辨的人类基因组重测序。我们还通过签名项目证明了邻接信息的显着价值，包括对胎儿基因组的第一次准确，非侵入性预测，以及对癌症基因组和表观基因组的第一次单倍型解析测序。在这个更新的应用程序中，我们建议将我们的重点缩小到我们的两个最有前途的方法，即接触概率映射（目标1）和邻接保持转座（目标2）的先进发展。然后，我们将正式评估这些方法的成本，性能和可扩展性，同时也寻求将它们相互整合，并与新兴的测序范式（目标3）。再加上短读段DNA测序的每碱基成本的适度下降，这些方法将使大基因组的染色体规模从头组装以及染色体规模单倍型分辨的人类基因组重测序成为可能，成本约为1000美元。

DESCRIPTION (provided by applicant): Even as new technologies continue to drive down the cost of DNA sequencing, we are in critical need of equivalently powerful methods informing long-range contiguity to support both de novo genome assembly and haplotype-resolved genome resequencing. With funding through this program, we have explored diverse approaches for low-cost, massively parallel capture of contiguity information. Our progress is substantial, and includes the development of a method for in situ library construction and optical sequencing, a method in which we exploit 'contact probability maps' to produce the first chromosome-scale de novo mammalian genome assemblies based exclusively on short reads, and a method that combines contiguity preserving transposition and combinatorial indexing for accurate, megabase-scale haplotype-resolved human genome resequencing. We have also demonstrated the remarkable value of contiguity information through signature projects, including the first accurate, non-invasive prediction of a fetal genome, and the first haplotype-resolved sequencing of a cancer genome and epigenome. In this renewal application, we propose to narrow our focus to the advanced development of our two most promising approaches, namely contact probability mapping (Aim 1) and contiguity preserving transposition (Aim 2). We will then formally evaluate these methods for cost, performance and scalability, while also seeking to integrate them with one another and with emerging sequencing paradigms (Aim 3). Coupled with a modest drop in the per-base cost of short read DNA sequencing, these methods will enable chromosome-scale de novo assembly of large genomes as well as chromosome-scale haplotype-resolved human genome resequencing for about $1,000.