Ultra high-throughput DNA synthesis via nano-optical conveyer belts

通过纳米光学传送带进行超高通量 DNA 合成

基本信息

批准号：
9379771
负责人：
Ronald Wayne Davis
金额：
$ 30.19万
依托单位：
STANFORD UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2017
资助国家：
美国
起止时间：
2017-08-09 至 2019-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9379771
关键词：
Acids Address Agriculture Automation Biology Caliber Cleaved cell Clinical DNA DNA Sequence DNA biosynthesis DNA sequencing Development Diploidy Disease Electrical Engineering Engravings Ethics Generations Genes Genetic Variation Genome Genomic medicine Genomics Guanine + Cytosine Composition Hazardous Waste Health Human Genome Human Genome Project Individual Industry Standard Investigation Laboratories Lasers Length Light Lighting Location Methods Nucleotides Oligonucleotides Optics Physiologic pulse Polystyrenes Production Property Reaction Reagent Recording of previous events Research Personnel Running Sampling Speed Surface Technology Temperature Testing Time To specify Writing base cost cost effective cost effectiveness gene function genetic variant genome editing interest melting nano nanoparticle novel optical traps phosphoramidite plasmonics precision medicine qubit social implication synthetic biology synthetic construct tool virtual

项目摘要

PROJECT SUMMARY The ability to understand genome biology and the consequences of genetic variation on individual health depends heavily on technologies that allow researchers to read and write DNA. Sequencing technologies have recently seen dramatic improvements that have made personal genomes affordable for virtually any laboratory and even directly available to consumers. Yet the corresponding DNA synthesis technologies lag far behind these developments, causing a major hindrance in synthetic biology efforts to study genes, variants, and genomes of interest by synthesizing them. This proposal aims to develop a novel DNA synthesis technology to address the greatest challenge faced by current platforms: maintaining sufficient accuracy for precision applications and throughput for large-scale applications while remaining cost-effective for accessibility. To achieve this, the traditional phosphoramidite method of synthesizing DNA oligonucleotides will be adapted onto nanoparticular beads, which will be moved through droplets containing synthesis reagents along a plasmonic surface array. This `conveyer belt' will be optically controlled via C-shaped engravings (CSEs) that concentrate light from below, serving as optical traps. In this way, the beads and reagent droplets can be individually, rapidly transported to specific optical traps in multiple lanes simply by changing the illumination wavelengths, allowing millions of unique oligonucleotides to be synthesized simultaneously on a single array. By tailoring the reagent droplet size and concentration depending on synthesis scale, the method will be optimized to target the entire bead surface, maximize yield, and eliminate excess reagent usage. Quality will be assessed by testing synthesis of diverse DNA sequences. The main advantages of this novel DNA synthesis platform will include: 1) faster reactions (cycle time 45 sec), 2) lower error rate due to decreased acid exposure (<1:1000), 3) high yield (>5 attomoles/bead), 4) increased length of oligonucleotides (>300 bases) due to cleaner synthesis, 6) significantly less hazardous waste production, 7) generation of >25 million unique oligonucleotide sequences in a single run that can be individually isolated for downstream applications, and 8) a cost of $0.0000001/base, two orders of magnitude less than the least expensive method currently available. A DNA synthesis technology with these properties will enable unprecedented genomic investigations, allowing researchers to test the functional and clinical impact of thousands of genes and genetic variants.

项目概要了解基因组生物学以及遗传变异对个人健康影响的能力很大程度上依赖于研究人员能够读写 DNA 的技术。测序技术最近看到了巨大的进步，使个人基因组几乎任何人都可以负担得起实验室甚至直接提供给消费者。但相应的DNA合成技术却滞后远远落后于这些发展，对合成生物学研究基因的努力造成重大阻碍，通过合成它们来获得变体和感兴趣的基因组。该提案旨在开发一种新型 DNA 合成方法技术来解决当前平台面临的最大挑战：保持足够的精度精密应用和大规模应用的吞吐量，同时保持成本效益可达性。为了实现这一点，合成DNA寡核苷酸的传统亚磷酰胺方法将适应纳米颗粒，该颗粒将通过含有合成物的液滴移动沿着等离子体表面阵列的试剂。该“传送带”将通过 C 形进行光学控制雕刻（CSE）从下方聚集光线，充当光陷阱。这样，珠子和只需通过改变照明波长，可以合成数百万种独特的寡核苷酸同时在单个阵列上。通过根据不同情况定制试剂液滴大小和浓度合成规模，该方法将被优化以针对整个珠子表面，最大限度地提高产量，并消除试剂使用过量。将通过测试不同 DNA 序列的合成来评估质量。主要这种新型 DNA 合成平台的优点包括：1) 反应更快（循环时间 45 秒），2) 更低由于酸暴露减少而导致错误率 (<1:1000)，3) 高产量（>5 attomoles/珠），4) 增加长度由于更清洁的合成，寡核苷酸（> 300 个碱基），6) 危险废物的产生显着减少，7) 单次运行生成超过 2500 万个独特的寡核苷酸序列，可以单独分离下游应用，以及 8) 成本为 0.0000001 美元/基地，比最低成本低两个数量级目前可用的方法昂贵。具有这些特性的 DNA 合成技术将使前所未有的基因组研究，使研究人员能够测试其功能和临床影响数以千计的基因和遗传变异。