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Droplet barcoding for massively parallel single-molecule deep sequencing

Author

Listed:
  • Freeman Lan

    (California Institute for Quantitative Biosciences (QB3), University of California
    UC Berkeley - UCSF Bioengineering Graduate program, University of California)

  • John R. Haliburton

    (California Institute for Quantitative Biosciences (QB3), University of California
    Integrative Program in Quantitative Biology (iPQB) Biophysics Graduate program, University of California)

  • Aaron Yuan

    (California Institute for Quantitative Biosciences (QB3), University of California
    University of California)

  • Adam R. Abate

    (California Institute for Quantitative Biosciences (QB3), University of California
    UC Berkeley - UCSF Bioengineering Graduate program, University of California
    Integrative Program in Quantitative Biology (iPQB) Biophysics Graduate program, University of California
    Present address: Department of Bioengineering and Therapeutic Sciences, University of California, San Francisco, 1700 4th Street, San Francisco, California 94158, USA.)

Abstract

The ability to accurately sequence long DNA molecules is important across biology, but existing sequencers are limited in read length and accuracy. Here, we demonstrate a method to leverage short-read sequencing to obtain long and accurate reads. Using droplet microfluidics, we isolate, amplify, fragment and barcode single DNA molecules in aqueous picolitre droplets, allowing the full-length molecules to be sequenced with multi-fold coverage using short-read sequencing. We show that this approach can provide accurate sequences of up to 10 kb, allowing us to identify rare mutations below the detection limit of conventional sequencing and directly link them into haplotypes. This barcoding methodology can be a powerful tool in sequencing heterogeneous populations such as viruses.

Suggested Citation

  • Freeman Lan & John R. Haliburton & Aaron Yuan & Adam R. Abate, 2016. "Droplet barcoding for massively parallel single-molecule deep sequencing," Nature Communications, Nature, vol. 7(1), pages 1-10, September.
    • Handle: RePEc:nat:natcom:v:7:y:2016:i:1:d:10.1038_ncomms11784
    DOI: 10.1038/ncomms11784
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