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Virtual microfluidics for digital quantification and single-cell sequencing

Nature Methods volume 13, pages 759762 (2016) | Download Citation

Abstract

We have developed hydrogel-based virtual microfluidics as a simple and robust alternative to complex engineered microfluidic systems for the compartmentalization of nucleic acid amplification reactions. We applied in-gel digital multiple displacement amplification (dMDA) to purified DNA templates, cultured bacterial cells and human microbiome samples in the virtual microfluidics system, and demonstrated whole-genome sequencing of single-cell MDA products with excellent coverage uniformity and markedly reduced chimerism compared with products of liquid MDA reactions.

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Acknowledgements

The authors thank S. Kim, D. Feldman and A. Kulesa for advice on bioinformatics, microscopy and image analysis; N. Ranu and the Hung lab (Broad Institute) for bacterial samples; L. Griffith, G. Lagoudas, J. Borrajo and L. Morinishi for helpful discussions; and members of the Griffith lab (MIT), especially C. Chopko, J. Valdez and H. Lee, for hydrogel expertise. This work was supported in part by a Lawrence Summers Fellowship from the Broad Institute (L.X.), a Career Award at the Scientific Interface from the Burroughs Welcome Fund (P.C.B.) and grants from the Center for Microbiome Informatics and Therapeutics at MIT; a National Human Genome Research Institute, grant number U54HG003067, to the Broad Institute; the Center for Environmental Health Sciences at MIT and the Fijian Ministry of Health. The Broad Institute and MIT may seek to commercialize aspects of this work, and related applications for intellectual property have been filed.

Author information

Affiliations

  1. Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA.

    • Liyi Xu
    • , Ilana L Brito
    • , Eric J Alm
    •  & Paul C Blainey
  2. The Broad Institute of MIT and Harvard, Cambridge, Massachusetts, USA.

    • Liyi Xu
    • , Ilana L Brito
    • , Eric J Alm
    •  & Paul C Blainey
  3. The Center for Microbiome Informatics and Therapeutics, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA.

    • Ilana L Brito
    •  & Eric J Alm

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Contributions

P.C.B. and L.X. conceived the concept for this study. L.X. designed and implemented experiments and conducted data analysis. I.L.B. and E.J.A. provided Fiji microbiome samples. I.L.B. conducted analysis of gut microbe data. L.X., P.C.B. and I.L.B. wrote and all authors approved the manuscript.

Competing interests

The Broad Institute and MIT may seek to commercialize aspects of this work and related applications for intellectual property have been filed.

Corresponding author

Correspondence to Paul C Blainey.

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    Supplementary Figures 1–8, Supplementary Tables 1–8 and Supplementary Notes 1–5

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    Features of 117 FijiCOMP single-cell assemblies.

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    Supplementary software for data analysis

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DOI

https://doi.org/10.1038/nmeth.3955

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