Genome-scale De Novo DNA Synthesis on Electronic p-Chips with Sorting

电子 p 芯片上的基因组规模 De Novo DNA 分选合成

• 批准号: 9353538
• 负责人: WLODEK MANDECKI
• 金额: $ 7.14万
• 依托单位: PHARMASEQ, INC.
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-09-07 至 2017-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9353538
• 关键词: Award; Chemistry; Chromosomes; Cloning; Combinatorial Synthesis; Computers; Custom; DNA; DNA Sequence; DNA biosynthesis; Diagnostic; Electronics; Enzymes; Funding; Gene-Modified; Genes; Genome; Genomics; Geometry; Goals; Grant; Health; Large-Scale Sequencing; Lasers; Light; Methods; Microfluidics; Molecular Genetics; Oligonucleotides; Optics; Phase; Polymers; Positioning Attribute; Price; Process; Production; Reaction; Reading; Research; Scientist; Signal Transduction; Small Business Innovation Research Grant; Solid; Sorting - Cell Movement; Speed; System; Technology; Time; Vaccine Design; Work; base; combinatorial; cost; design; drug discovery; genetic variant; genome-wide; innovation; instrument; instrumentation; plasmid DNA; prototype; radio frequency; synthetic construct; synthetic genomics; tool

 DESCRIPTION (provided by applicant): The purpose of the present project is to develop and validate a new method for synthesizing large number of DNA fragments, oligodeoxynucleotides. The method, which is called "sequence-directed DNA synthesis", involves the use of small electronic circuits, "p-Chips", each of which transmits a unique ID number via RF when illuminated with laser light. In a massively parallel format, oligonucleotides are synthesized using p-Chips as the solid phase. The p-Chip ID provides the ability to associate a given DNA sequence with that particular p-Chip as synthesis proceeds. A key part of the system is a high-speed instrument, the sorter, that reads the ID of each p-Chip and then sorts p-Chips into the appropriate reaction vessel based on the p-Chip ID, thus the specific DNA sequence. The system, when completed, will be able to generate synthetic DNA fragments in very short times and at costs that are much lower than current methods. The work plan for the project is to initially, in Phase I, demonstrate the ability to synthesize DNA on currently available 0.5 mm p-Chips and to show feasibility of sorting. Following this, in Phase II, will shift to a new p-Chip design, each of which is one thousandth the volume of the 0.5 mm p-Chip. The main goals will be to build the sorter for these ultra-small chips, and to formulate a working method for large-scale sequence-directed combinatorial DNA synthesis. We will also demonstrate applicability of the method by synthesizing a large number of oligonucleotides on p-Chips and creating a 10 kb long gene fragment. We will obtain the DNA sequence error rate upon cloning and identify the best methods for gene assembly and construction of long segments of chromosomes. The prospect of rapidly producing DNA at low prices has profound implications in a number of fields related to synthetic genomics where new tools are needed. This includes the design of modified genes for enzymes, energy production, vaccine designs and many other applications.

 描述(申请人提供)：本项目的目的是开发和验证一种合成大量DNA片段的新方法--寡聚脱氧核苷酸。这种被称为“序列导向的DNA合成”的方法涉及到使用小型电子电路，即“p芯片”，每个芯片在激光照射下通过射频传输一个唯一的ID号。在大规模并行格式中，使用p-芯片作为固相合成寡核苷酸。P-Chip ID提供了在合成进行时将给定DNA序列与该特定p-Chip相关联的能力。该系统的一个关键部分是一个高速仪器，即分类器，它读取每个p芯片的ID，然后根据p芯片ID将p芯片分类到适当的反应容器中，从而获得特定的DNA序列。该系统建成后，将能够在非常短的时间内产生合成DNA片段，成本远远低于目前的方法。该项目的工作计划是，在第一阶段，初步展示在现有的0.5 mm p芯片上合成DNA的能力，并展示分选的可行性。随后，在第二阶段，将转向新的p-Chip设计，每种设计的体积都是0.5 mm p-Chip的千分之一。主要目标将是为这些超小芯片建造分类器，并制定一种大规模序列定向组合DNA合成的工作方法。我们还将通过在p芯片上合成大量寡核苷酸并创建10kb长的基因片段来证明该方法的适用性。我们将获得克隆时的DNA序列错误率，并确定基因组装和构建染色体长片段的最佳方法。以低价快速生产DNA的前景对合成基因组学相关的许多领域具有深远的影响，这些领域需要新的工具。这包括用于酶、能源生产、疫苗设计和许多其他应用的修饰基因的设计。