A Microfluidic DNA Library Preparation Platform for Next-Generation Sequencing

Next-generation sequencing (NGS) is emerging as a powerful tool for elucidating genetic information for a wide range of applications. Unfortunately, the surging popularity of NGS has not yet been accompanied by an improvement in automated techniques for preparing formatted sequencing libraries. To address this challenge, we have developed a prototype microfluidic system for preparing sequencer-ready DNA libraries for analysis by Illumina sequencing. Our system combines droplet-based digital microfluidic (DMF) sample handling with peripheral modules to create a fully-integrated, sample-in library-out platform. In this report, we use our automated system to prepare NGS libraries from samples of human and bacterial genomic DNA. E. coli libraries prepared on-device from 5 ng of total DNA yielded excellent sequence coverage over the entire bacterial genome, with >99% alignment to the reference genome, even genome coverage, and good quality scores. Furthermore, we produced a de novo assembly on a previously unsequenced multi-drug resistant Klebsiella pneumoniae strain BAA-2146 (KpnNDM). The new method described here is fast, robust, scalable, and automated. Our device for library preparation will assist in the integration of NGS technology into a wide variety of laboratories, including small research laboratories and clinical laboratories.

下一代测序（Next-generation Sequencing，NGS）正逐渐成为阐明多类应用场景下遗传信息的强有力工具。遗憾的是，NGS的普及度迅猛提升的同时，用于制备合规测序文库的自动化技术却并未得到相应发展。为应对这一挑战，我们研发了一款微流控原型系统，可用于制备可直接上机进行Illumina测序分析的DNA文库。本系统将液滴式数字微流控（Droplet-based Digital Microfluidic，DMF）样本处理技术与外围模块相结合，打造出一套全集成的样本输入-文库输出平台。本研究中，我们利用这套自动化系统从人类及细菌基因组DNA样本中制备了NGS文库。从5纳克总DNA中在本设备上制备的大肠杆菌（E. coli）文库，可在整个细菌基因组上实现优异的序列覆盖度，与参考基因组的比对率超过99%，基因组覆盖均匀且质量评分良好。此外，我们对一株此前未被测序的多重耐药肺炎克雷伯菌菌株BAA-2146（KpnNDM）完成了从头组装（de novo assembly）。本文所述的新方法具备快速、稳健、可扩展且自动化的优势。我们的文库制备设备将助力NGS技术融入各类实验室，包括小型科研实验室与临床实验室。