A Microfluidic-Based SNP Genotyping Method for Hereditary Hearing-Loss Detection
A Microfluidic-Based SNP Genotyping Method for Hereditary Hearing-Loss Detection
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基于微流控的 SNP 基因分型方法用于遗传性听力损失检测
DOI：
10.1021/acs.analchem.9b00652
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发表时间：
2019-05-07
影响因子：
7.4
通讯作者：
Xu，Youchun
中科院分区：
文献类型：
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作者：
Lu，Ying;Chen，Shan;Xu，Youchun
单核苷酸多态性（SNP）的基因分型是许多遗传性疾病（包括遗传性耳聋）分析的先决条件。然而，现有的SNP检测方法存在检测周期长、操作繁琐和携带污染风险高的问题。为了解决这些挑战，本研究构建了一种用于快速高效SNP基因分型的微流控芯片，该芯片将样品分成许多独立的腔室进行竞争等位基因特异性PCR。采用这种策略，可以很容易地实现多重检测，并从根本上克服了建立多重PCR的挑战。与SNaPshot和MassArray相比，该方法可以在2 h内完全密封地完成整个检测过程。在对该芯片的基本性能进行评估后，应用该芯片筛选了15个突变，包括SNPs和InDels（插入-缺失标记），可以覆盖中国80%以上的遗传性耳聋病例。利用该芯片对40多份临床样本进行SNP分型，结果与桑格测序结果一致，表明该芯片在遗传病检测中的实用性和应用潜力。
The genotyping of SNPs (single nucleotide polymorphisms) is a prerequisite for the analysis of many genetic diseases, including hereditary hearing-loss. However, the existing methods for SNP detection suffer from a long detection period, tedious operation, and a high risk of carryover contamination. To address these challenges, a microfluidic chip is constructed for rapid and efficient SNP genotyping by dividing the sample into many independent chambers for Kompetitive Allele Specific PCR in this study. Using this strategy, multiple detection can be easily accomplished and the challenge for the establishment of multiplex PCR is fundamentally overcome. The entire detection can be finished within 2 h in a fully sealed manner with this method, which is quite simple compared to SNaPshot and MassArray. After assessment of the basic performance, this chip was applied to screen 15 mutations, including SNPs and InDels (insertion-deletion markers), that can cover more than 80% of cases of hereditary hearing-loss in China. Over 40 clinical samples were analyzed with this microfluidic chip for SNP genotyping, and the results are consistent with that obtained by Sanger sequencing, demonstrating its practicability and potential in the application of genetic disease detection.