Miniaturized Integrated Genotyping Microsystems

小型化集成基因分型微系统

• 批准号: 6943017
• 负责人: RICHARD A MATHIES
• 金额: $ 24.7万
• 依托单位: UNIVERSITY OF CALIFORNIA BERKELEY
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-09-01 至 2009-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6943017
• 关键词: biomedical equipment development; biopsy; capillary electrophoresis; genetic markers; genotype; high throughput technology; miniature biomedical equipment; mitochondrial DNA; molecular oncology; molecular pathology; neoplasm /cancer diagnosis; neoplasm /cancer invasiveness; polymerase chain reaction; portable biomedical equipment

DESCRIPTION (provided by applicant): The objective of this grant is to develop novel microfabricated genetic analysis microsystems and associated methods that can be used for high-performance analysis of cancer genotypes in the research, discovery and/or diagnostic settings. Initial work will focus on the refinement of the apparatus, reagents and methods needed to apply Polymorphism Ratio Sequencing (PRS) to the high-throughput genetic analysis of mitochondrial DNA variations in tumor tissue using conventional capillary array electrophoresis. We will optimize the labeling and pooling methods and develop convenient PRS data analysis software. Then, Johns Hopkins University (JHU) scientists will be trained to perform PRS at UCB. Finally, we will transition the technique to JHU for its routine high-throughput application. Second, we will design, construct and evaluate a fully integrated mitochondrial PRS chip. This wafer scale device takes RCA (rolling circle amplification) prepared mitochondrial DNA and parses the template into 96 individual DNA sequencing modules, including extension reactors and CE separation channels, to produce an entire mitochondrial PRS analysis in under 1 hr. Once this system is developed, a second-generation version will be constructed and then used at JHU for high-throughput analyses. Third, we will develop a fully integrated microdevice to perform SNP and other genetic typing from genomic DNA. This device will accept purified genomic DNA as the input and will parse the individual sample to 96 different PCR reactors for multiplex allele specific amplification and analysis of polymorphisms or cancer markers. This microdevice will permit genetic typing from small quantities of DNA and has the advantage of fully integrating a large portion of the important sample preparation process thereby providing low-cost, high-throughput genotyping of tumor samples. Finally, we will develop a portable genotyping device for real-time analysis of informative mitochondiral or genomic DNA variations or diagnostic markers. This system will be valuable (i) for point-of-care genetic analysis to identify the presence of cancer markers and/or to monitor possible recurrence and (ii) for performing real-time molecular pathology of tissue samples to determine the extent of cancer invasion.

描述（由申请人提供）： 该赠款的目的是开发新型的微生物遗传分析微生系统和相关方法，这些方法可用于研究，发现和/或诊断环境中的癌症基因型的高性能分析。最初的工作将集中在使用常规毛细管阵列电泳的肿瘤组织中线粒体DNA变异的高通量遗传分析中，将多态性比率测序（PRS）应用于高通量遗传分析所需的试剂和方法。我们将优化标签和汇总方法，并开发方便的PRS数据分析软件。然后，约翰·霍普金斯大学（JHU）科学家将接受培训，可以在UCB进行PRS。最后，我们将把技术转移到JHU，以供其常规的高通量应用程序。其次，我们将设计，构建和评估完全集成的线粒体PRS芯片。该晶圆刻度设备需要RCA（滚动圆扩增）制备的线粒体DNA，并将模板解析为96个单独的DNA测序模块，包括延伸反应器和CE分离通道，以在1小时以下的1小时内产生整个线粒体PRS分析。一旦开发了该系统，将构建第二代版本，然后在JHU中用于高通量分析。第三，我们将开发一个完全集成的微电位，以执行基因组DNA的SNP和其他遗传键入。该设备将接受纯化的基因组DNA作为输入，并将单个样品解析为96个不同的PCR反应器，以进行多重等位基因特异性扩增和分析多态性或癌症标志物。该微电位将允许少量DNA的遗传分类，并具有充分整合重要样品制备过程中大部分部分的优势，从而提供低成本的高通量基因分型的肿瘤样品。最后，我们将开发一种便携式基因分型设备，用于实时分析线粒体或基因组DNA变异或诊断标记。该系统将是有价值的（i）用于确定癌症标志物和/或监测可能复发的存在以及（ii）进行组织样本实时分子病理以确定癌症侵袭程度的潜在复发和（ii）的有价值（i）。