Commercialization of a low-cost user-friendly DNA preparation kit that produces chromosome-span contiguity from conventional short-read sequencing​​ for a wide range of applications

生产染色体跨度的低成本、用户友好的DNA制备试剂盒的商业化 与传统短读测序的邻接性适用于广泛的应用 ARIMA基因组学 7.项目摘要/摘要 超过90%的下一代测序(NGS)是通过Illumina短读测序仪进行测序的。这是 因为它的成本效益和更快的周转时间。然而，短读测序技术 丢失关键的邻接性信息，在重新组装基因组和重建母体基因组方面受到限制 二倍体基因组的父本单倍型。邻接性信息对于理解遗传学很有价值 这对人类健康和疾病至关重要，因此对于提高精确度和个性化医疗至关重要。长- Read技术(如太平洋生物科学)只能达到百万级的染色体邻接性，但却是5-7倍 比Illumina Short-Read更贵，限制了它的使用。DNA制备的最新进展可以保存 与Illumina短读测序兼容的远程信息。这些“人工合成”的长阅读(或 SLR)方法可以改进短读技术，具有远距离连续性和短读经济性。 然而，最大的SLR邻接性只有100Mb人类平均染色体邻接性的1-5%。至 构建多兆位连续，SLR方法需要基因组DNA(GDNA)片段&gt；100-150kb，但 获得长的gDNA片段是具有挑战性的，这限制了当前的SLR方法。ARIMA基因组学已经 将HIC技术重新用于基于SLR的DNA制备试剂盒，以保持染色体跨度的连续性 以及短读兼容的单倍型阶段化。ARIMA的关键技术创新是，而不是使用 纯化的gDNA，我们的方法利用了自然保存在三维空间中的长时间连续信息 (3D)基因组在细胞中的组织，因为3D信息是远程信息。通过捕获3D基因组 信息，ARIMA的Contigo试剂盒重建了染色体跨度信息。来自ARIMA的Contigo套件 因此与短线经济具有完全的连续性。Contigo套件价格实惠，不需要 深奥的试剂、其他技能或设备，并与现有的短期工作流程兼容，以提供 用户可轻松采用。此外，Contigo是保存3D基因组信息的唯一试剂盒 解读非编码功能基因组学。Contigo试剂盒有可能成为 新一代测序在基因分型、分阶段、等位基因特异性表达等方面的广泛应用 以及阶段性基因组的表观遗传学、结构变异分析、元基因组学和3D基因组学。在这 提议我们将Contigo技术商业化，通过提高对广泛细胞输入和 基因组覆盖的一致性，并实现96井兼容。此外，我们将为制造而设计， 降低成本并改进采购，确保在实际运输和存储条件下的强劲性能， 改善用户体验，最后，与领域专家Key一起对Contigo工具包的性能进行基准测试 意见领袖和强大的用户来展示商业可行性。

Commercialization of a low-cost user-friendly DNA preparation kit that produces chromosome-span contiguity from conventional short-read sequencing for a wide range of applications Arima Genomics 7. Project Summary/Abstract Over 90% of Next-Generation Sequencing (NGS) is sequenced via Illumina short-read sequencers. This is because of its cost-effectiveness and faster turn-around times. However, short-read sequencing technologies lose critical contiguity information and are limited in assembling genomes de novo and reconstructing maternal and paternal haplotypes of diploid genomes. Contiguity information is valuable for understanding the genetics of human health and disease, and therefore critical for advancing precision and personalized medicine. Long- read technologies (e.g. Pacific Biosciences) only reach megabase-scale chromosomal contiguity, but are 5-7X more expensive than Illumina short-read, limiting its use. Recent advances in DNA preparation can preserve long-range information that is compatible with Illumina short-read sequencing. These “synthetic” long-reads (or SLR) methods can improve short-read technologies with, long-range contiguity and short-read economy. However, the maximal SLR contiguity is only 1-5% the contiguity a 100Mb average human chromosome. To construct multi-megabase contiguity, SLR methods require genomic DNA (gDNA) fragments >100-150Kb, but obtaining long gDNA fragments is challenging and this limits current SLR methods. Arima Genomics has repurposed HiC technology as an SLR-based DNA preparation kit that preserves chromosome-span contiguity and haplotype phasing that is short-read compatible. Arima key technical innovation is, rather than using purified gDNA, our method leverages the long-contiguity information preserved naturally in the 3-dimensional (3D) organization of genomes in cells, as 3D information is long-range information. By capturing 3D genome information, Arima's Contigo kit reconstructs chromosome-span information. The Contigo kit from Arima therefore has complete contiguity with short-read economy. The Contigo kit is affordable, does not require esoteric reagents, additional skills or equipment, and is compatible with exiting short-read workflows, to offer easy user adoption. In addition, Contigo is the only kit to preserve 3D genomic information essential to interpreting non-coding functional genomics. The Contigo kit has the potential to serve as a new standard in next-generation sequencing with broad applications in genotyping, phasing, allele-specific gene expression and epigenetics on phased genomes, structural variation analyses, metagenomics and 3D genomics. In this proposal we will commercialize the Contigo technology by improving sensitivity to wide range of cell-inputs and uniformity in genome coverage, and enabling 96-well compatibility. In addition, we will design for manufacture, reduce cost and improve sourcing, assure robust performance in real-world shipping and storage conditions, improve user-experience, and finally, benchmark performance of the Contigo kit with domain experts, key opinion leaders, and power users to demonstrate commercial viability.