Research ArticleNANOTECHNOLOGY

Visible/near-infrared subdiffraction imaging reveals the stochastic nature of DNA walkers

See allHide authors and affiliations

Science Advances  20 Jan 2017:
Vol. 3, no. 1, e1601600
DOI: 10.1126/sciadv.1601600
  • Fig. 1 DNA walker system design and walking mechanism.

    (A) Schematic of a DNA walker system on a coverslip. The coverslip surface is passivated by the combination of hydrophobic coating [dichlorodimethylsilane (DDS)] and surfactant (Tween 20). Biotinylated bovine serum albumin (biotin-BSA) is embedded into the passivation layer as an anchor point for the nanotube track. Carbon nanotubes functionalized with biotinylated RNA and RNA fuel strands bind to the surface through streptavidin/biotin interaction. The DNAzyme-decorated CdTe/CdS QD conjugates with the immobilized nanotube track through base pairing. (B) AFM image of a DNA-QD conjugated with an RNA-SWCNT. The striation observed along the tube axis is a result of RNA wrapping. (C) A single-turnover event of the enzymatic walking mechanism.

  • Fig. 2 Experimental setup and imaging scheme.

    (A) Excitation from two laser lines are configured as reflected light epi-illumination. Sample emissions are collected by an objective lens and separated into two light paths (VIS and NIR) for imaging with corresponding cameras. a.u., arbitrary units. (B) (i and ii) The PSF of a QD is fitted using a Gaussian function. The averaged localizations from 10 images form a final position estimation with ~20 nm precision. (iii and iv) Intensity fluctuation of the pixel labeled #1 in the NIR nanotube image. (v to vii) The image of isolated intensity fluctuation spot is obtained by constructing a difference image of successive frames. (viii) The isolated PSFs are localized to reconstruct the subdiffraction nanotube image. (ix) The localized VIS and NIR images are registered to the same coordinates to produce the overlaid image. The QD position is indicated by a green arrow. The scale bars are 500 nm, unless otherwise indicated.

  • Fig. 3 Images of a DZ7 DNAzyme walker moving along an RNA-SWCNT track.

    (A) Raw and (B) subdiffraction images of a DZ7 DNAzyme walker in PBS containing 10 mM Mg2+ and 20 mM DTT. The walker travels over 200 nm in 10 min. Scale bars, 500 nm. (C) Walking trajectory plotted in a 400-nm × 400-nm grid for better visualization of the travel distance.

  • Fig. 4 Quantitative evaluation of a 10–23 DNAzyme walker.

    (A) Three representative displacements of the walker (blue scatter). The smoothed displacement from 10 successive localizations is presented in blue curves. Data from a control experiment (0 mM Mg2+) are shown in black scatter and black curve. (B) Velocity distribution obtained from all displacement curves. Gaussian fitted histogram yields 〈v〉 = 7.8 ± 0.37 nm min−1. (C) MSD plot of the walker. The time-averaged MSD is shown in thin black lines, and the ensemble-averaged data are plotted in black scatter. A power-law function is fitted to the experimental data with a scaling exponent of 1.7. The first 10 scatter points are linear-fitted (blue line) to better visualize the deviation of the walker MSD from diffusive motion. (D) Experimental and simulated mean displacement variance. The simulated results with a different number of rate-limiting steps (n) are plotted as thin curves in different colors: n = 1, black; n = 2, red; n = 3, blue. The experimental data are plotted in scatter points and linear-fitted (blue line).

  • Table 1 Walker randomness evaluation.
    MethodsIntermediate reactionsDesigned rObserved r
    Simulation111.03 ± 0.01
    Simulation20.50.49 ± 0.01
    Simulation30.330.32 ± 0.01
    Stage111.02 ± 0.12
    Stage20.50.67 ± 0.13
    Stage30.330.39 ± 0.08
    DNA walker30.34 ± 0.15

Supplementary Materials

  • Supplementary material for this article is available at http://advances.sciencemag.org/cgi/content/full/3/1/e1601600/DC1

    Supplementary Methods

    fig. S1. DNA-QD size distribution and RNA-SWCNT length fractionation.

    fig. S2. Optical properties of DNA-QDs.

    fig. S3. Imaging surface characterization.

    fig. S4. Characterization of the localization and drift correction precision.

    fig. S5. Images of a 10-23 DNAzyme walker moving along an RNA-SWCNT track.

    fig. S6. Displacement plots of the walking experiment data sets.

    fig. S7. Numerical simulations of DNA walker translocation.

    table S1. Sequence information about DNA walker and RNA fuel strands.

    movie S1. Raw and subdiffraction images showing a DZ7 DNAzyme walker moving along an RNA-SWCNT track.

    References (4246)

  • Supplementary Materials

    This PDF file includes:

    • Supplementary Methods
    • fig. S1. DNA-QD size distribution and RNA-SWCNT length fractionation.
    • fig. S2. Optical properties of DNA-QDs.
    • fig. S3. Imaging surface characterization.
    • fig. S4. Characterization of the localization and drift correction precision.
    • fig. S5. Images of a 10-23 DNAzyme walker moving along an RNA-SWCNT track.
    • fig. S6. Displacement plots of the walking experiment data sets.
    • fig. S7. Numerical simulations of DNA walker translocation.
    • table S1. Sequence information about DNA walker and RNA fuel strands.
    • movie S1. Raw and subdiffraction images showing a DZ7 DNAzyme walker moving along an RNA-SWCNT track.
    • References (42–46)

    Download PDF

    Download Movie S1

    Files in this Data Supplement:

Navigate This Article