Massively Parallel Contiguity Mapping

描述（由申请人提供）：即使新技术不断降低DNA测序的成本，我们也迫切需要同样强大的方法来通知远程邻近，以支持从头基因组组装和单倍型解析基因组重测序。在这个项目的资助下，我们探索了多种低成本、大规模并行捕获邻近信息的方法。我们的进展是实质性的，包括原位文库构建和光学测序方法的开发，一种利用“接触概率图”产生第一个基于短读的染色体尺度的从头哺乳动物基因组组装的方法，以及一种结合了保持相邻转位和组合索引的方法，用于精确的，超大规模的单倍型解决人类基因组重测序。我们还通过签名项目证明了邻近信息的显著价值，包括第一个准确的、非侵入性的胎儿基因组预测，以及第一个单倍型解决癌症基因组和表观基因组测序。在这个更新应用中，我们建议将我们的重点缩小到我们两种最有前途的方法的先进发展上，即接触概率映射（Aim 1）和保持相邻的转置（Aim 2）。然后，我们将正式评估这些方法的成本、性能和可扩展性，同时也寻求将它们彼此整合，并与新兴的测序范例整合（目标3）。再加上短读DNA测序每碱基成本的适度下降，这些方法将使大型基因组的染色体规模从头组装以及染色体规模单倍型解析的人类基因组重测序成本约为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.