Development of Fluorescent Detector for DNA Sequencing

描述（由申请人提供）： 人类序列变异最常见的形式是单核苷酸多态性（SNP），它可以影响蛋白质功能，基因的正确加工或影响基因表达的正常水平。我们建议开发一种新的荧光检测方法，应用于高通量识别信息SNP，这可能会导致更准确的诊断遗传性疾病，更好的预后风险易感性，或零星突变的识别。该技术被称为脉冲多线激励或PME。PME技术具有两个潜在的优点，这可以显著提高荧光灵敏度：（1）基因组测定中所有荧光团的最佳激发和（2）“色盲”检测，其收集更多的光。该技术与目前最先进的DNA测序仪器有很大的不同，后者的特点是单源激发和色散，用于DNA序列鉴定。为了测试PME用于多色荧光检测的可行性，我们提出了单个毛细管试验板的构建，详细的检测限实验以评估灵敏度，以及用于SNP的DNA测序的重建实验。 PME技术的成功实施将广泛应用于临床诊断、法医学和一般测序方法的常规使用，并将具有针对大多数人群的靶向序列变异测定的能力、灵活性和便携性。

DESCRIPTION (provided by applicant): The most commonly observed form of human sequence variation is single nucleotide polymorphisms (SNPs), which can affect protein function, proper processing of genes or affect the normal level of gene expression. We propose the development of a novel approach to fluorescence detection with application for high-throughput identification of informative SNPs, which could lead to more accurate diagnosis of inherited disease, better prognosis of risk susceptibilities, or identification of sporadic mutations. The proposed technology is called Pulse-Multiline Excitation or PME. The PME technology has two potential advantages, which could significantly increase fluorescence sensitivity: (1) optimal excitation of all fluorophores in the genomic assay and (2) "color-blind" detection, which collects considerably more light. This technology differs significantly from the current state-of-the-art DNA sequencing instrumentation, which features single source excitation and color dispersion for DNA sequence identification. To test the feasibility of PME for multi-color fluorescence detection, we propose the construction of a single capillary breadboard, detailed limit of detection experiments to assess sensitivity, and reconstruction experiments for DNA sequencing of SNPs. Successful implementation of the PME technology will have broad application for routine usage in clinical diagnostics, forensics, and general sequencing methodologies and will have the capability, flexibility, and portability of targeted sequence variation assays for a large majority of the population.