A Microfluidic Device for Multiplex Single-Nucleotide Polymorphism Genotyping.
A Microfluidic Device for Multiplex Single-Nucleotide Polymorphism Genotyping.
复制标题
用于多重单核苷酸多态性基因分型的微流体装置。
DOI：
10.1039/c3ra44091e
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发表时间：
2014
期刊：
影响因子：
3.9
通讯作者：
Lin,Qiao
中科院分区：
文献类型：
--
作者：
Zhu,Jing;Qiu,Chunmei;Palla,Mirkó;Nguyen,ThaiHuu;Russo,JamesJ;Ju,Jingyue;Lin,Qiao
单核苷酸多态性（SNPs）是最丰富的遗传变异类型;它们提供了个体的遗传指纹，对于发现遗传生物标志物至关重要。SNPs的准确检测对于疾病的预防、诊断和预后以及预测患者的药物反应和临床结局具有重要意义。然而，传统的SNP基因分型方法仍然受到准确性不足或劳动、时间和资源密集型程序的限制。微流控技术已越来越多地用于提高效率;然而，目前可用的微流控基因分型系统在准确性、灵敏度、通量和多路复用能力方面仍然存在缺点。为了应对这些挑战，我们开发了一种多步骤SNP基因分型微流控装置，该装置在温控芯片上进行SNP特异性引物的单碱基延伸和延伸产物的固相纯化。这些产品可通过基质辅助激光解吸/电离飞行时间质谱（MALDI-TOF MS）立即检测，从而鉴定目标基因座的等位基因。该集成装置能够实现高效和自动化操作，同时保持MS提供的高准确度和灵敏度。通过对合成模板上的4个基因座进行快速、准确和同时检测，验证了多重基因分型能力。微流控装置具有执行自动、准确、定量和高通量分析的潜力，涵盖生物和临床研究、药物开发和法医学中的广泛应用。
Single-nucleotide polymorphisms (SNPs) are the most abundant type of genetic variations; they provide the genetic fingerprint of individuals and are essential for genetic biomarker discoveries. Accurate detection of SNPs is of great significance for disease prevention, diagnosis and prognosis, and for prediction of drug response and clinical outcomes in patients. Nevertheless, conventional SNP genotyping methods are still limited by insufficient accuracy or labor-, time-, and resource-intensive procedures. Microfluidics has been increasingly utilized to improve efficiency; however, the currently available microfluidic genotyping systems still have shortcomings in accuracy, sensitivity, throughput and multiplexing capability. To address these challenges, we developed a multi-step SNP genotyping microfluidic device, which performs single-base extension of SNP specific primers and solid-phase purification of the extension products on a temperature-controlled chip. The products are ready for immediate detection by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), providing identification of the alleles at the target loci. The integrated device enables efficient and automated operation, while maintaining the high accuracy and sensitivity provided by MS. The multiplex genotyping capability was validated by performing rapid, accurate and simultaneous detection of 4 loci on a synthetic template. The microfluidic device has the potential to perform automatic, accurate, quantitative and high-throughput assays covering a broad spectrum of applications in biological and clinical research, drug development and forensics.