Preparation of Next-Generation Sequencing Libraries Using Nextera™ Technology: Simultaneous DNA Fragmentation and Adaptor Tagging by In Vitro Transposition
DNA library preparation is a common entry point and bottleneck for next-generation sequencing. Current methods generally consist of distinct steps that often involve significant sample loss and hands-on time: DNA fragmentation, end-polishing, and adaptor-ligation. In vitro transposition with Nextera™ Transposomes simultaneously fragments and covalently tags the target DNA, thereby combining these three distinct steps into a single reaction. Platform-specific sequencing adaptors can be added, and the sample can be enriched and bar-coded using limited-cycle PCR to prepare di-tagged DNA fragment libraries. Nextera technology offers a streamlined, efficient, and high-throughput method for generating bar-coded libraries compatible with multiple next-generation sequencing platforms.
Key wordsNext-generation sequencing DNA library preparation Roche/454 Illumina/Solexa Nextera
The author would like to thank Jay Shendure and Hilary Morrison for their work characterizing the Illumina-compatible and Roche-compatible libraries and Haiying Grunenwald for technical advice and assistance during the development of this method.
Illumina and Solexa are registered trademarks of Illumina Inc., San Diego, CA. Tween is a registered trademark of ICI Americas Inc., Wilmington, DE. 454 and GS FLX are trademarks of Roche, Nutley, NJ. DNA Clean & Concentrator and Zymo-Spin are trademarks of Zymo Research., Orange, CA. Nextera and Tagmentation are trademarks of EPICENTRE, Madison, WI. Patent applications assigned to EPICENTRE and by US Patent Nos. 5,965,443, and 6,437,109; European Patent No. 0927258, and related patents and patent applications, exclusively licensed to EPICENTRE, cover Nextera™ Products.