Nanostructured Magnetic Disks for Wide Range Size Selection in NGS Library Preparation and PCR Purification

用于 NGS 文库制备和 PCR 纯化中宽范围尺寸选择的纳米结构磁盘

• 批准号: 10212466
• 负责人: Kelvin Liu
• 金额: $ 41.07万
• 依托单位: CIRCULOMICS, INC.
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-08-18 至 2021-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10212466
• 关键词: Biochemical Reaction; Caliber; DNA; Data; Development; Diagnostic tests; Disease; Genetic; Genome; Human Resources; Length; Libraries; Magnetism; Methods; Molecular Weight; Nanostructures; National Human Genome Research Institute; Preparation; Protocols documentation; Reaction; Recovery; Sampling; Silicon Dioxide; Structure; Testing; Time; Translations; Validation; cost; density; improved; novel diagnostics; pan-genome; particle; prevent; reference genome; therapeutic target

Project Summary We proposed to develop Nanobind reaction purification and size selection methods for long-read sequencing library preparation. These methods use our Nanobind magnetic disks to purify enzymatic reactions and to enhance read lengths by removing short DNA during long-read sequencing library preparation. Nanobind disks are small 1-5 mm diameter magnetic disks covered with a high density of micro- and nanostructured silica. Compared to magnetic particles, Nanobind disks are extraordinarily gentle, preventing DNA shearing and enabling high efficiency recovery of DNA surpassing megabase in length. We currently sell DNA extraction kits that enable isolation of ultra high molecular weight (UHMW) megabase-sized DNA from a wide variety of sample types. To evaluate how gentle the Nanobind reaction purifications are, we tested them in an ultra-long sequencing protocol capable of generating 100 kb – 1+ Mb length reads. This method is capable of generating extremely high read length N50 in the 100 – 200 kb range and 10 times more megabase reads than previous ultra long methods. This supplement will allow additional development and sequencing tests on the Nanobind ultra long sequencing protocols. Successful validation of the method on PromethION will lead to a significant enhancement in data throughput and corresponding in reduction of cost, time, and manpower, enabling an immediate impact via translation into projects such as the NHGRI pan-genome project and a long-term impact by dramatically reducing the cost of ultra long sequencing.

项目摘要 我们建议开发纳米素反应纯化和尺寸选择方法，以进行长阅读测序 图书馆准备。这些方法使用我们的纳米素磁盘来纯化酶促反应和 通过在长阅读测序库制备过程中删除短DNA来增强读取长度。纳米磁盘 是小1-5毫米直径的磁盘，上面覆盖着高密度的微型和纳米结构二氧化硅。 与磁性颗粒相比，纳米粒磁盘非常温和，可防止DNA剪切和 能够高度恢复DNA的长度超过巨核。我们目前出售DNA提取套件 可以从各种样品中分离出超高分子量（UHMW）巨型Babase大小的DNA 类型。为了评估纳米素反应净化的温和程度，我们在超长中测试了它们 测序协议能够生成100 kb - 1+ MB长度读取。此方法能够生成 在100 - 200 kb范围内读取长度N50的读数极高，巨型Babase读数是以前的10倍 超长方法。这种补充剂将允许对纳米宾的其他开发和测序测试 超长测序方案。成功验证该方法在promethion上将导致重大 增强数据吞吐量和减少成本，时间和人力的对应关系，使 通过转化为NHGRI PAN-GENOME项目和长期影响等项目的直接影响 通过大大降低超长测序的成本。