High-throughput automated microfluidic sample preparation for accurate microbial genomics.
High-throughput automated microfluidic sample preparation for accurate microbial genomics.
复制标题
DOI：
10.1038/ncomms13919
复制
发表时间：
2017-01-27
影响因子：
16.6
通讯作者：
Blainey PC
中科院分区：
文献类型：
--
作者：
Kim S;De Jonghe J;Kulesa AB;Feldman D;Vatanen T;Bhattacharyya RP;Berdy B;Gomez J;Nolan J;Epstein S;Blainey PC
低成本的鸟枪式DNA测序正在改变微生物科学。测序仪器是如此有效，以至于样品准备现在是关键的限制因素。在这里，我们介绍了一个微流控样品制备平台，它集成了细胞中的关键步骤，可以为多达96个样品测序库样品制备，并在保持或提高数据质量的同时将DNA输入需求减少100倍。我们演示的通用微体系结构支持具有任意数量的反应和清理或捕获步骤的工作流。通过减少样本数量要求，我们实现了低输入(∼10,000个细胞)全基因组鸟枪法(WGS)对结核分枝杆菌和土壤微菌落的测序，获得了优异的结果。我们还利用提高的吞吐量对∼400临床铜绿假单胞菌文库进行了测序，并展示了出色的单核苷酸多态检测性能，这解释了观察到的抗生素耐药性的表型。完全集成的芯片上实验室样品制备克服了技术障碍，使基因组学能够在许多基础研究和翻译应用中更广泛地部署。微生物基因组分析的鸟枪式DNA测序实验通常是不切实际的，因为需要最少的样本输入。在这里，作者开发了一种微流控样品制备平台，它可以将样品输入要求减少100倍，并能够从较少的细胞数量进行高通量测序。
Low-cost shotgun DNA sequencing is transforming the microbial sciences. Sequencing instruments are so effective that sample preparation is now the key limiting factor. Here, we introduce a microfluidic sample preparation platform that integrates the key steps in cells to sequence library sample preparation for up to 96 samples and reduces DNA input requirements 100-fold while maintaining or improving data quality. The general-purpose microarchitecture we demonstrate supports workflows with arbitrary numbers of reaction and clean-up or capture steps. By reducing the sample quantity requirements, we enabled low-input (∼10,000 cells) whole-genome shotgun (WGS) sequencing of Mycobacterium tuberculosis and soil micro-colonies with superior results. We also leveraged the enhanced throughput to sequence ∼400 clinical Pseudomonas aeruginosa libraries and demonstrate excellent single-nucleotide polymorphism detection performance that explained phenotypically observed antibiotic resistance. Fully-integrated lab-on-chip sample preparation overcomes technical barriers to enable broader deployment of genomics across many basic research and translational applications. Shotgun DNA sequencing experiments for microbial genomic analysis are often impractical due to minimum sample input requirements. Here the authors develop a microfluidic sample preparation platform that reduces sample input requirements 100-fold and enables high throughput sequencing from low numbers of cells.