Fully Streched Single DNA Molecules in a Nanofluidic Chip Show Large-Scale Structural Variation

Jonas Nyvold Pedersen, Rodolphe Marie, D. L. Bauer, Kristian Hagsted Rasmussen, Mohamed Adan Yusuf, E. V. Volpi, Anders Kristensen, K. U. Mir, Henrik Flyvbjerg

    Research output: Contribution to journalComment/debateResearchpeer-review

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    Abstract

    When stretching and imaging DNA molecules in nanofluidic devices, it is important to know the relation between the physical length as measured in the lab and the distance along the contour of the DNA. Here a single DNA molecule longer than 1 Mbp is loaded into a nanofluidic device consisting of two crossing nanoslits (85nm x 50 microns) connected to microchannels. An applied pressure creates a stagnation point at the crossing of the nanoslits. The drag force from the fluid stretches the DNA. We determine the degree of stretching of the molecule (i) without the use of markers, (ii) without knowing the contour length of the DNA, and (iii) without having the full DNA molecule inside the field-of-view. The analysis is based on the transverse motion of the DNA due its Brownian motion, i.e. the DNA's response to the thermal fluctuations of the liquid surrounding it. The parameter values obtained by fitting agree well with values we obtain from simplified modeling of the DNA as a cylinder in a parallel flow. Secondly, DNA molecules stained with the intercalating dye YOYO-1 are de- and renatured locally following a modified version of the protocol used in Ref. 1. The result is a melting pattern which reflects the local AT/GC-content. Single molecules are loaded into the chip and imaged. Due to the almost complete stretching of the DNA, structural variations in the size range from kbp to Mbp can be detected and quantified from the melting pattern alone.

    Original languageEnglish
    JournalBiophysical Journal
    Volume104
    Issue number2; SUPP/1
    Pages (from-to)175a
    ISSN0006-3495
    Publication statusPublished - 2013

    Cite this

    @article{2ebf2c9ffeee4a1a963b533e1be4f615,
    title = "Fully Streched Single DNA Molecules in a Nanofluidic Chip Show Large-Scale Structural Variation",
    abstract = "When stretching and imaging DNA molecules in nanofluidic devices, it is important to know the relation between the physical length as measured in the lab and the distance along the contour of the DNA. Here a single DNA molecule longer than 1 Mbp is loaded into a nanofluidic device consisting of two crossing nanoslits (85nm x 50 microns) connected to microchannels. An applied pressure creates a stagnation point at the crossing of the nanoslits. The drag force from the fluid stretches the DNA. We determine the degree of stretching of the molecule (i) without the use of markers, (ii) without knowing the contour length of the DNA, and (iii) without having the full DNA molecule inside the field-of-view. The analysis is based on the transverse motion of the DNA due its Brownian motion, i.e. the DNA's response to the thermal fluctuations of the liquid surrounding it. The parameter values obtained by fitting agree well with values we obtain from simplified modeling of the DNA as a cylinder in a parallel flow. Secondly, DNA molecules stained with the intercalating dye YOYO-1 are de- and renatured locally following a modified version of the protocol used in Ref. 1. The result is a melting pattern which reflects the local AT/GC-content. Single molecules are loaded into the chip and imaged. Due to the almost complete stretching of the DNA, structural variations in the size range from kbp to Mbp can be detected and quantified from the melting pattern alone.",
    author = "Pedersen, {Jonas Nyvold} and Rodolphe Marie and Bauer, {D. L.} and Rasmussen, {Kristian Hagsted} and Yusuf, {Mohamed Adan} and Volpi, {E. V.} and Anders Kristensen and Mir, {K. U.} and Henrik Flyvbjerg",
    year = "2013",
    language = "English",
    volume = "104",
    pages = "175a",
    journal = "Biophysical Journal",
    issn = "0006-3495",
    publisher = "Cell Press",
    number = "2; SUPP/1",

    }

    Fully Streched Single DNA Molecules in a Nanofluidic Chip Show Large-Scale Structural Variation. / Pedersen, Jonas Nyvold; Marie, Rodolphe ; Bauer, D. L.; Rasmussen, Kristian Hagsted; Yusuf, Mohamed Adan; Volpi, E. V.; Kristensen, Anders; Mir, K. U.; Flyvbjerg, Henrik.

    In: Biophysical Journal, Vol. 104, No. 2; SUPP/1, 2013, p. 175a.

    Research output: Contribution to journalComment/debateResearchpeer-review

    TY - JOUR

    T1 - Fully Streched Single DNA Molecules in a Nanofluidic Chip Show Large-Scale Structural Variation

    AU - Pedersen, Jonas Nyvold

    AU - Marie, Rodolphe

    AU - Bauer, D. L.

    AU - Rasmussen, Kristian Hagsted

    AU - Yusuf, Mohamed Adan

    AU - Volpi, E. V.

    AU - Kristensen, Anders

    AU - Mir, K. U.

    AU - Flyvbjerg, Henrik

    PY - 2013

    Y1 - 2013

    N2 - When stretching and imaging DNA molecules in nanofluidic devices, it is important to know the relation between the physical length as measured in the lab and the distance along the contour of the DNA. Here a single DNA molecule longer than 1 Mbp is loaded into a nanofluidic device consisting of two crossing nanoslits (85nm x 50 microns) connected to microchannels. An applied pressure creates a stagnation point at the crossing of the nanoslits. The drag force from the fluid stretches the DNA. We determine the degree of stretching of the molecule (i) without the use of markers, (ii) without knowing the contour length of the DNA, and (iii) without having the full DNA molecule inside the field-of-view. The analysis is based on the transverse motion of the DNA due its Brownian motion, i.e. the DNA's response to the thermal fluctuations of the liquid surrounding it. The parameter values obtained by fitting agree well with values we obtain from simplified modeling of the DNA as a cylinder in a parallel flow. Secondly, DNA molecules stained with the intercalating dye YOYO-1 are de- and renatured locally following a modified version of the protocol used in Ref. 1. The result is a melting pattern which reflects the local AT/GC-content. Single molecules are loaded into the chip and imaged. Due to the almost complete stretching of the DNA, structural variations in the size range from kbp to Mbp can be detected and quantified from the melting pattern alone.

    AB - When stretching and imaging DNA molecules in nanofluidic devices, it is important to know the relation between the physical length as measured in the lab and the distance along the contour of the DNA. Here a single DNA molecule longer than 1 Mbp is loaded into a nanofluidic device consisting of two crossing nanoslits (85nm x 50 microns) connected to microchannels. An applied pressure creates a stagnation point at the crossing of the nanoslits. The drag force from the fluid stretches the DNA. We determine the degree of stretching of the molecule (i) without the use of markers, (ii) without knowing the contour length of the DNA, and (iii) without having the full DNA molecule inside the field-of-view. The analysis is based on the transverse motion of the DNA due its Brownian motion, i.e. the DNA's response to the thermal fluctuations of the liquid surrounding it. The parameter values obtained by fitting agree well with values we obtain from simplified modeling of the DNA as a cylinder in a parallel flow. Secondly, DNA molecules stained with the intercalating dye YOYO-1 are de- and renatured locally following a modified version of the protocol used in Ref. 1. The result is a melting pattern which reflects the local AT/GC-content. Single molecules are loaded into the chip and imaged. Due to the almost complete stretching of the DNA, structural variations in the size range from kbp to Mbp can be detected and quantified from the melting pattern alone.

    M3 - Comment/debate

    VL - 104

    SP - 175a

    JO - Biophysical Journal

    JF - Biophysical Journal

    SN - 0006-3495

    IS - 2; SUPP/1

    ER -