A Parallel High-Throughput Pharmacogenetic Profiling Assay for Clinical Use

药物基因组学研究已经确定了许多基因，其SNP基因型或单倍型与不同的个体药物反应、其他代谢过程或疾病易感性相关。因此，快速准确地确定医学相关区域的基因型的能力，对于理解个人基因谱对这些过程的影响，以及对预测、预防和个性化医学的发展至关重要。Maxwell Sensors Inc.提议开发条形码蚀刻可测微珠（BEAM Bead）技术，该技术将数字条形码珠和分子化学相结合，用于药物遗传SNP基因分型分析，可用于个体的高通量分子分析。数字BEAM微珠是顺磁性的，通过光刻技术制造，具有数千个识别码，可以与核酸，蛋白质或其他小分子功能化，在均质或非均质介质中进行大型多路检测。使用BEAM珠进行基因分型具有灵活性、高通量、易于制造、低成本批量生产以及在小批量的一次反应中完成的优点。在项目的第一阶段，我们成功地(1)制造了10位BEAM磁珠，允许1,024（=210）个唯一代码，使其成为世界上复用率最高的条形码磁珠；(2)设计并构建了微流体传感器和荧光检测器，逐个解码检测荧光；(3)建立P450 （CYP2A7）、KCNA5和ITPase的多重PCR，并进行头杂交；(4)利用多重PCR扩增序列评估SNP基因分型分析。二期工作将集中于优化临床应用的BEAM头平台，设计和集成BEAM系统，设计标准和定制SNP面板，用于与药物反应相关的SNP基因分型，以及开发用于靶基因扩增和SBE杂交的多重PCR。

DESCRIPTION (provided by applicant): A Parallel High Throughput Pharmacogenetic Profiling Assay for Clinical Use Pharmacogenomic studies have already identified a number of genes whose SNP genotypes or haplotypes correlate with different individual drug responses, other metabolic processes or disease susceptibility. Thus the ability to determine genotypes quickly and accurately for medically relevant regions is critical to understanding the effects of an individual's genetic profile on these processes, and to the development of predictive, preventative and personalized medicine. Maxwell Sensors Inc. proposes to develop Barcode Etched Assayable Micro Bead (BEAM bead) technology, which combines digital barcoded beads and molecular chemistry for a pharmacogenetic SNP genotyping assay, which can be used for the high throughput molecular profiling of individuals. The digital BEAM beads are paramagnetic and are fabricated by photolithography to have thousands of identification codes and can be functionalized with nucleic acids, proteins or other small molecules to carry out large multiplexed assays in homogeneous or heterogeneous media. Genotyping using BEAM beads offers the advantages of flexibility, high throughput, easy fabrication, low cost mass production, and all in one reaction in small volumes. During Phase I of the project, we have successfully (1) fabricated 10 digit BEAM beads, allowing 1,024 (=210) unique codes, making it the most highly multiplexed barcode magnetic bead in the world; (2) designed and constructed a microfluidic transducer and fluorescence detector to decode and detect fluorescence one by one; (3) developed multiplex PCR for P450 (CYP2A7), KCNA5 and ITPase, and bead hybridization; and (4) evaluated the SNP genotyping assay using multiplex PCR amplified sequences. Phase II work will focus on optimization of the BEAM bead platform for clinical applications, design and integration of BEAM system, design of standard and custom SNP panels for genotyping SNPs relevant to drug responses, and development of multiplex PCR for target gene amplification and SBE hybridization.