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Real-time selective sequencing using nanopore technology

Abstract

The Oxford Nanopore Technologies MinION sequencer enables the selection of specific DNA molecules for sequencing by reversing the driving voltage across individual nanopores. To directly select molecules for sequencing, we used dynamic time warping to match reads to reference sequences. We demonstrate our open-source Read Until software in real-time selective sequencing of regions within small genomes, individual amplicon enrichment and normalization of an amplicon set.

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Figure 1: Enrichment of lambda genome target regions by direct sequence fragment selection using Read Until.
Figure 2: Selective sequencing of specific amplicons from a library using Read Until.
Figure 3: Normalization of amplicon sequencing effort in a library using Read Until.

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European Nucleotide Archive

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European Nucleotide Archive

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Acknowledgements

This work was supported by BBSRC TRDF award BB/M020061/1. The authors thank T. Evans for comments and discussion on the manuscript and figures, N. Loman and J. Quick for helpful discussions, advice and motivation and M. Blythe for discussions. We also thank ONT for access to the Read Until API and helpful discussions and support.

Author information

Affiliations

Authors

Contributions

M.L. conceived and designed the project, wrote the code, ran sequencing, performed analysis and wrote the manuscript. M.S. carried out analysis and wrote and contributed to code and contributed to the manuscript. S.M. generated libraries, ran sequencing and contributed to the manuscript.

Corresponding author

Correspondence to Matthew Loose.

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Competing interests

M.L. is a member of the MinION access program (MAP) and has received free-of-charge flowcells and kits for nanopore sequencing and travel and accommodation expenses to speak at Oxford Nanopore Technologies conferences.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–11 and Supplementary Tables 1–4 (PDF 8749 kb)

Supplementary Software

Read Until code and accessory files (ZIP 2016 kb)

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Loose, M., Malla, S. & Stout, M. Real-time selective sequencing using nanopore technology. Nat Methods 13, 751–754 (2016). https://doi.org/10.1038/nmeth.3930

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