New in-Situ Method for On-chip DNA Oligomer Synthesis

芯片上 DNA 寡聚物原位合成新方法

• 批准号: 6444218
• 负责人: ROBERT C HAUSHALTER
• 金额: $ 12.14万
• 依托单位: PARALLEL SYNTHESIS TECHNOLOGIES, INC.
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-05-15 至 2003-04-14
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6444218
• 关键词: biotechnology; in situ hybridization; liquid chromatography mass spectrometry; method development; microarray technology; nucleic acid chemical synthesis; nucleic acid sequence; oligonucleotides; phosphamide; silicon

The use of DNA arrays for numerous applications has expanded rapidly in recent years. These arrays are made primarily using photolabile ligands as blocking groups in an array format or by spotting DNA from organisms or a DNA synthesizer. We propose an on-chip, high-density DNA oligomer growth method that will use the standard phosphoramidite chemistry on a proprietary chip design. The key to the technology will be a chip that contains an array of <500um grids regions micromachined directly into a silicon wafer using Si microfabrication techniques. The grid regions are micromeshes fabricated by a wet chemical etching method giving grid mesh element sizes selectable from 10-100um. The growing oligonucleotides are attached to the SiO2 surface of the grid. This array of grids is placed over a second wafer that has <500um holes in registry with, and underneath, the micromesh areas. The ability of a DNA synthesizer to force equilibrium to completion by flowing reagents through the stationary support, and to thoroughly wash each sample, is duplicated on this chip by placing the nucleotide synthesis chemicals on the mesh and then drawing them through the mesh/hole regions of each chip, thereby simulating the flow through geometry of a DNA synthesizer. PROPOSED COMMERCIAL APPLICATION: If feasibility is demonstrated, the economics of silicon microfabrication and mass production will allow the fabrication of custom DNA microarrays by this method to compete very effectively with the current technology. Since only a commercial liquid handling robot and the proposed chips would be necessary to automate this system, the cost reduction could greatly accelerate the use of microarray technology in biology and genetic investigations.

近年来，DNA阵列在许多应用中的使用迅速扩大。这些阵列主要使用光不稳定配体作为阵列格式中的阻断基团或通过从生物体或DNA合成仪点样DNA来制备。我们提出了一种芯片上，高密度DNA寡聚体生长方法，将使用标准的亚磷酰胺化学的专有芯片设计。该技术的关键将是一个芯片，其中包含一个阵列的<500微米网格区域微加工直接进入硅晶片使用硅微加工技术。网格区域是通过湿法化学蚀刻方法制造的微网格，从而给出可从10- 100 μ m选择的网格网格单元尺寸。生长的寡核苷酸附着在网格的SiO2表面。将该网格阵列放置在第二晶片上，该第二晶片具有与微网区域对准并在其下方的<500 μ m的孔。通过将核苷酸合成化学品放置在网格上，然后将它们穿过每个芯片的网格/孔区域，从而模拟DNA合成仪的流动几何形状，在该芯片上复制了DNA合成仪通过使试剂流过固定支持物而强制平衡完成以及彻底洗涤每个样品的能力。拟定商业应用：如果可行性得到证明，硅微加工和大规模生产的经济性将允许通过这种方法制造定制DNA微阵列，以非常有效地与当前技术竞争。由于只有一个商业化的液体处理机器人和拟议的芯片将是必要的自动化这个系统，降低成本可以大大加快微阵列技术在生物学和遗传研究中的使用。