Sequencing of human genomes extracted from single cancer cells isolated in a valveless microfluidic device

Rodolphe Marie*, Marie Pødenphant, Kamila Koprowska, Loic Baerlocher, Roland C. M. Vulders, Jennifer Wilding, Neil Ashley, Simon J. McGowan, Dianne van Strijp, Freek van Hemert, Tom Olesen, Niels Agersnap, Brian Bilenberg, Celine Sabatel, Julien Schira, Anders Kristensen, Walter Bodmer, Pieter J. van der Zaag, Kalim U. Mir

*Corresponding author for this work

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    Abstract

    Sequencing the genomes of individual cells enables the direct determination of genetic heterogeneity amongst cells within a population. We have developed an injection-moulded valveless microfluidic device in which single cells from colorectal cancer derived cell lines (LS174T, LS180 and RKO) and fresh colorectal tumors have been individually trapped, their genomes extracted and prepared for sequencing using multiple displacement amplification (MDA). Ninety nine percent of the DNA sequences obtained mapped to a reference human genome, indicating that there was effectively no contamination of these samples from non-human sources. In addition, most of the reads are correctly paired, with a low percentage of singletons (0.17 +/- 0.06%) and we obtain genome coverages approaching 90%. To achieve this high quality, our device design and process shows that amplification can be conducted in microliter volumes as long as the lysis is in sub-nanoliter volumes. Our data thus demonstrates that high quality whole genome sequencing of single cells can be achieved using a relatively simple, inexpensive and scalable device. Detection of genetic heterogeneity at the single cell level, as we have demonstrated for freshly obtained single cancer cells, could soon become available as a clinical tool to precisely match treatment with the properties of a patient's own tumor.
    Original languageEnglish
    JournalLab on a Chip
    Volume18
    Issue number13
    Pages (from-to)1891-1902
    Number of pages12
    ISSN1473-0197
    DOIs
    Publication statusPublished - 2018

    Bibliographical note

    Open Access Article. This article is licensed under a Creative Commons Attribution 3.0 Unported Licence.

    Cite this

    Marie, R., Pødenphant, M., Koprowska, K., Baerlocher, L., Vulders, R. C. M., Wilding, J., ... Mir, K. U. (2018). Sequencing of human genomes extracted from single cancer cells isolated in a valveless microfluidic device. Lab on a Chip, 18(13), 1891-1902. https://doi.org/10.1039/c8lc00169c
    Marie, Rodolphe ; Pødenphant, Marie ; Koprowska, Kamila ; Baerlocher, Loic ; Vulders, Roland C. M. ; Wilding, Jennifer ; Ashley, Neil ; McGowan, Simon J. ; van Strijp, Dianne ; van Hemert, Freek ; Olesen, Tom ; Agersnap, Niels ; Bilenberg, Brian ; Sabatel, Celine ; Schira, Julien ; Kristensen, Anders ; Bodmer, Walter ; van der Zaag, Pieter J. ; Mir, Kalim U. / Sequencing of human genomes extracted from single cancer cells isolated in a valveless microfluidic device. In: Lab on a Chip. 2018 ; Vol. 18, No. 13. pp. 1891-1902.
    @article{5b3367e12d5748eda2d47588a4543354,
    title = "Sequencing of human genomes extracted from single cancer cells isolated in a valveless microfluidic device",
    abstract = "Sequencing the genomes of individual cells enables the direct determination of genetic heterogeneity amongst cells within a population. We have developed an injection-moulded valveless microfluidic device in which single cells from colorectal cancer derived cell lines (LS174T, LS180 and RKO) and fresh colorectal tumors have been individually trapped, their genomes extracted and prepared for sequencing using multiple displacement amplification (MDA). Ninety nine percent of the DNA sequences obtained mapped to a reference human genome, indicating that there was effectively no contamination of these samples from non-human sources. In addition, most of the reads are correctly paired, with a low percentage of singletons (0.17 +/- 0.06{\%}) and we obtain genome coverages approaching 90{\%}. To achieve this high quality, our device design and process shows that amplification can be conducted in microliter volumes as long as the lysis is in sub-nanoliter volumes. Our data thus demonstrates that high quality whole genome sequencing of single cells can be achieved using a relatively simple, inexpensive and scalable device. Detection of genetic heterogeneity at the single cell level, as we have demonstrated for freshly obtained single cancer cells, could soon become available as a clinical tool to precisely match treatment with the properties of a patient's own tumor.",
    author = "Rodolphe Marie and Marie P{\o}denphant and Kamila Koprowska and Loic Baerlocher and Vulders, {Roland C. M.} and Jennifer Wilding and Neil Ashley and McGowan, {Simon J.} and {van Strijp}, Dianne and {van Hemert}, Freek and Tom Olesen and Niels Agersnap and Brian Bilenberg and Celine Sabatel and Julien Schira and Anders Kristensen and Walter Bodmer and {van der Zaag}, {Pieter J.} and Mir, {Kalim U.}",
    note = "Open Access Article. This article is licensed under a Creative Commons Attribution 3.0 Unported Licence.",
    year = "2018",
    doi = "10.1039/c8lc00169c",
    language = "English",
    volume = "18",
    pages = "1891--1902",
    journal = "Lab on a Chip",
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    Marie, R, Pødenphant, M, Koprowska, K, Baerlocher, L, Vulders, RCM, Wilding, J, Ashley, N, McGowan, SJ, van Strijp, D, van Hemert, F, Olesen, T, Agersnap, N, Bilenberg, B, Sabatel, C, Schira, J, Kristensen, A, Bodmer, W, van der Zaag, PJ & Mir, KU 2018, 'Sequencing of human genomes extracted from single cancer cells isolated in a valveless microfluidic device', Lab on a Chip, vol. 18, no. 13, pp. 1891-1902. https://doi.org/10.1039/c8lc00169c

    Sequencing of human genomes extracted from single cancer cells isolated in a valveless microfluidic device. / Marie, Rodolphe ; Pødenphant, Marie; Koprowska, Kamila; Baerlocher, Loic; Vulders, Roland C. M.; Wilding, Jennifer; Ashley, Neil; McGowan, Simon J.; van Strijp, Dianne; van Hemert, Freek; Olesen, Tom; Agersnap, Niels; Bilenberg, Brian; Sabatel, Celine; Schira, Julien; Kristensen, Anders; Bodmer, Walter; van der Zaag, Pieter J.; Mir, Kalim U.

    In: Lab on a Chip, Vol. 18, No. 13, 2018, p. 1891-1902.

    Research output: Contribution to journalJournal articleResearchpeer-review

    TY - JOUR

    T1 - Sequencing of human genomes extracted from single cancer cells isolated in a valveless microfluidic device

    AU - Marie, Rodolphe

    AU - Pødenphant, Marie

    AU - Koprowska, Kamila

    AU - Baerlocher, Loic

    AU - Vulders, Roland C. M.

    AU - Wilding, Jennifer

    AU - Ashley, Neil

    AU - McGowan, Simon J.

    AU - van Strijp, Dianne

    AU - van Hemert, Freek

    AU - Olesen, Tom

    AU - Agersnap, Niels

    AU - Bilenberg, Brian

    AU - Sabatel, Celine

    AU - Schira, Julien

    AU - Kristensen, Anders

    AU - Bodmer, Walter

    AU - van der Zaag, Pieter J.

    AU - Mir, Kalim U.

    N1 - Open Access Article. This article is licensed under a Creative Commons Attribution 3.0 Unported Licence.

    PY - 2018

    Y1 - 2018

    N2 - Sequencing the genomes of individual cells enables the direct determination of genetic heterogeneity amongst cells within a population. We have developed an injection-moulded valveless microfluidic device in which single cells from colorectal cancer derived cell lines (LS174T, LS180 and RKO) and fresh colorectal tumors have been individually trapped, their genomes extracted and prepared for sequencing using multiple displacement amplification (MDA). Ninety nine percent of the DNA sequences obtained mapped to a reference human genome, indicating that there was effectively no contamination of these samples from non-human sources. In addition, most of the reads are correctly paired, with a low percentage of singletons (0.17 +/- 0.06%) and we obtain genome coverages approaching 90%. To achieve this high quality, our device design and process shows that amplification can be conducted in microliter volumes as long as the lysis is in sub-nanoliter volumes. Our data thus demonstrates that high quality whole genome sequencing of single cells can be achieved using a relatively simple, inexpensive and scalable device. Detection of genetic heterogeneity at the single cell level, as we have demonstrated for freshly obtained single cancer cells, could soon become available as a clinical tool to precisely match treatment with the properties of a patient's own tumor.

    AB - Sequencing the genomes of individual cells enables the direct determination of genetic heterogeneity amongst cells within a population. We have developed an injection-moulded valveless microfluidic device in which single cells from colorectal cancer derived cell lines (LS174T, LS180 and RKO) and fresh colorectal tumors have been individually trapped, their genomes extracted and prepared for sequencing using multiple displacement amplification (MDA). Ninety nine percent of the DNA sequences obtained mapped to a reference human genome, indicating that there was effectively no contamination of these samples from non-human sources. In addition, most of the reads are correctly paired, with a low percentage of singletons (0.17 +/- 0.06%) and we obtain genome coverages approaching 90%. To achieve this high quality, our device design and process shows that amplification can be conducted in microliter volumes as long as the lysis is in sub-nanoliter volumes. Our data thus demonstrates that high quality whole genome sequencing of single cells can be achieved using a relatively simple, inexpensive and scalable device. Detection of genetic heterogeneity at the single cell level, as we have demonstrated for freshly obtained single cancer cells, could soon become available as a clinical tool to precisely match treatment with the properties of a patient's own tumor.

    U2 - 10.1039/c8lc00169c

    DO - 10.1039/c8lc00169c

    M3 - Journal article

    VL - 18

    SP - 1891

    EP - 1902

    JO - Lab on a Chip

    JF - Lab on a Chip

    SN - 1473-0197

    IS - 13

    ER -