Loading and selective release of cargo in DNA nanotubes with longitudinal variation

Journal name:
Nature Chemistry
Volume:
2,
Pages:
319–328
Year published:
DOI:
doi:10.1038/nchem.575
Received
Accepted
Published online

Abstract

Nanotubes hold promise for a number of biological and materials applications because of their high aspect ratio and encapsulation potential. A particularly attractive goal is to access nanotubes that exert well-defined control over their cargo, such as selective encapsulation, precise positioning of the guests along the nanotube length and triggered release of this cargo in response to specific external stimuli. Here, we report the construction of DNA nanotubes with longitudinal variation and alternating larger and smaller capsules along the tube length. Size-selective encapsulation of gold nanoparticles into the large capsules of these tubes leads to ‘nanopeapod’ particle lines with positioning of the particles 65 nm apart. These nanotubes can then be opened when specific DNA strands are added to release their particle cargo spontaneously. This approach could lead to new applications of self-assembled nanotubes, such as in the precise organization of one-dimensional nanomaterials, gene-triggered selective delivery of drugs and biological sensing.

At a glance

Figures

  1. Construction of triangular DNA nanotubes 3, 4, 5 and 6.
    Figure 1: Construction of triangular DNA nanotubes 3, 4, 5 and 6.

    a, Nine dsLS of appropriate sequence were designed to assemble pairs of small triangles 1 and large triangles 2 to give triangular DNA nanotubes 3 with longitudinal variation and alternating larger (14 nm) and smaller (7 nm) capsules along the tube length. b, Size-selective encapsulation of AuNPs (gold spheres) gave nanotubes 4 that encapsulated lines of nanoparticles in the large capsules, with precise positioning of the particles 65 nm apart. c, Two linking strands that connected together triangles 2 into the large capsule were modified to contain an eight-base overhang (ES1). Assembly of triangles 1 and 2, the appropriate linking strands and the new linking strands (ES1) in the presence of 15 nm AuNPs resulted in DNA nanotubes 5. Selective opening of DNA nanotubes 5 with specific added DNA eraser strands ES1′, which were fully complementary to ES1, led to nanotubes 6 in their single-stranded form that spontaneously released their particle cargo.

  2. Construction of triangular rungs 1 and 2.
    Figure 2: Construction of triangular rungs 1 and 2.

    a, DNA of the appropriate length, sequence, and number of organic branch points 13 is synthesized using automated solid-phase methods to generate linear DNA strands, which can subsequently be cleaved from the solid support and used to build triangles, as shown in (b) and (c). The blue, green and orange strands denote different sequences. b, (i) The linear strand 7 is subsequently hybridized into a triangle using a complementary DNA strand (red), and then (ii) chemically ligated using cyanogen bromide to yield the single-stranded DNA template 8, after denaturation. (v) Native polyacrylamide gel electrophoresis (PAGE) analysis reveals the clean templated closure of a triangle (lane 3) using the linear strand 7 (lane 2) and the complementary template strand (lane 1). (vi) Denaturing PAGE analysis reveals the generation of the chemically ligated cyclic triangle 8 as a single other band of relatively retarded electrophoretic mobility (lane 2). (iii–iv, vii) The single-stranded and cyclic template 8 (lane 1) is sequentially titrated with the complementary strands CS1–CS3 (lane 2–4, respectively), and with the rigidifying strands RS1–RS3 to quantitatively generate a fully assembled triangular rung 1 (lane 5). (c) (i) Two linear strands 10a and 10b are hybridized using a complementary DNA template, and then (ii) chemically ligated using cyanogen bromide to yield the linear single-stranded 11, after denaturation. (v) Native PAGE analysis reveals the clean templated hybridization product (lane 4) of linear strands 10a and 10b (lane 2 and lane 3) and the complementary template strand (lane 1). (vi) Denaturing PAGE analysis reveals the generation of chemically ligated product 11 as a single band of relatively retarded electrophoretic mobility when the assemblies of 10a and 10b (lane 1 and 2) are chemically ligated using cyanogen bromide (lane 3). (iii,iv,vii) The strand 11 (lane 1) is sequentially titrated with the complementary strands CS1′CS3′ (lane 2–4, respectively), and with strands RS1–RS3 to quantitatively generate a fully triangular rung 2.

  3. Characterization of DNA nanotubes 3.
    Figure 3: Characterization of DNA nanotubes 3.

    a–c, The construction of alternating large–small DNA nanotubes 3 from triangular rungs 1 and 2 (see Fig. 1a) resulted in well-defined, one-dimensional DNA assemblies that extended over 10 µm, as shown by the AFM height image (a) and phase image (b); the AFM phase image of DNA nanotubes of uniform size longitudinally is shown for comparison (c) (scale bars = 1 µm). d, TEM images of Pt–C replicas revealed the corrugated architecture of these nanotubes, with a distance of ∼65 nm between the repeat features lengthwise, as well as an average diameter ratio of the large to small capsules that is very close to the theoretically calculated value of 2.0 (oval areas) (scale bars = 100 nm). e, For comparison, TEM image of Pt–C replicas of DNA nanotubes of uniform size longitudinally (scale bar = 100 nm). f, Confocal fluorescence intensity images show nanotubes 3 to be long and stable in solution at room temperature. The right column illustrates the counts per millisecond per pixel (scale bar = 10 µm).

  4. AuNP encapsulation into alternating large–small triangular-shaped DNA nanotubes 4.
    Figure 4: AuNP encapsulation into alternating large–small triangular-shaped DNA nanotubes 4.

    a,b, High-resolution AFM images clearly show encapsulation of the AuNPs within the triangular-shaped DNA nanotubes 4 (generated from triangular rungs 1 and 2 (see Fig. 1b)) into a ‘peapod’ architecture. c, High-resolution AFM images show the filled tubes bundle next to each other completely symmetrically, with their encapsulated particles next to each other laterally. d,e, Cryo-EM (d) and TEM (e) images show the linear pattern of AuNPs. f, TEM statistical analysis of the distances between AuNPs in each line gave ∼63 or ∼120 nm, which is consistent with the position of the particles in the large capsules of these nanotubes (yellow oval line). However, control experiments I, II and III (see Fig. 5a–c, respectively) mostly showed aggregation of the AuNPs (black oval line). ac, Scale bars = 250 nm, de, scale bars = 100 nm.

  5. Control experiments.
    Figure 5: Control experiments.

    a–c, The obtained nanoparticles and nanotubes observed by AFM (i) and TEM (ii), with the corresponding control procedure (iii) (a–c correspond to I–III in Fig. 4f, respectively). a, AuNPs of 10 nm average diameter were added to the nanotube components and nanotube 3 was assembled in their presence; no encapsulation was observed. b, 15 nm AuNPs were added to the pre-formed DNA nanotube 3; no ordering of these particles into lines was observed, which confirmed that the observed particle lines arose from encapsulated particles, rather than from particles bound to the exterior of the DNA nanotubes. c, 15 nm AuNPs were added only to the small triangular rungs 1, and these rungs were linked into a DNA nanotube with regular 7 nm cavities; no encapsulation and ordering of the particles was observed. Scale bars = 1 µm for AFM (i) and 100 nm for TEM (ii) images.

  6. Characterization of the DNA nanotubes 5 for selective release of AuNPs in response to specific external DNA eraser strands ES1′.
    Figure 6: Characterization of the DNA nanotubes 5 for selective release of AuNPs in response to specific external DNA eraser strands ES1′.

    a,b, AFM and TEM images of nanotubes 5 (produced from 1, 2 and new modified linking strands ES, with an eight-base overhang in the presence of 15 nm AuNPs (see Fig. 1c)) and 6 (produced when a fully complementary eraser DNA strand ES1′ was added (see Fig. 1c)). a,i AFM images showed stiff DNA nanotubes 5, with the AuNPs loading within the large cavities of the tubes, and organized into nanopeapod lines (scale bar = 250 nm); this was also confirmed by TEM images (ii) (scale bar = 100 nm). b, When the eraser DNA strand ES1′ was added, AFM images (i) showed flexible nanotubes 6 with reduced persistence length, consistent with their partially single-stranded character (scale bar = 250 nm). No encapsulated AuNP architecture was observed in the TEM images (ii) (scale bar = 100 nm) after the addition of the eraser strand ES1′. c, TEM statistical analysis revealed loss of ordering of the nanoparticles, and the distances were no longer consistent with their positioning within the nanotubes.

  7. Construction of DNA nanotubes 13a and 13b, and ultraviolet–visible spectra of 13a.
    Figure 7: Construction of DNA nanotubes 13a and 13b, and ultraviolet–visible spectra of 13a.

    a, Two linking strands that connect together triangles 2 into large nanotubes were modified to contain an eight-base overhang. Assembly of triangles 2 and the appropriate linking strand plus the new linking strands ES3 in the presence of 20 nm AuNP resulted in DNA nanotubes 13a with uniformly encapsulated and closely spaced AuNPs. Selective opening of the DNA nanotubes 13a with specific added DNA strands ES3′ led to nanotubes 13b in their single-stranded form that spontaneously released their particle cargo. TEM images showed the line pattern of AuNPs in close proximity in nanotubes 13a and the loss of ordering of AuNPs 15 minutes after the addition of strand ES3′ (scale bars = 200 nm). b, Ultraviolet–visible spectra of nanotubes 13a recorded at various times after addition of ‘eraser’ strand ES3′ at room temperature from zero to 15 minutes (arrows show increase and decrease in intensity with time).

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Author information

Affiliations

  1. Department of Chemistry, McGill University, 801 Sherbrooke Street West, Montreal, QC H3A 2K6, Canada

    • Pik Kwan Lo,
    • Pierre Karam,
    • Faisal A. Aldaye,
    • Christopher K. McLaughlin,
    • Graham D. Hamblin,
    • Gonzalo Cosa &
    • Hanadi F. Sleiman

Contributions

All authors discussed the results and commented on the manuscript. H.F.S. conceived and designed the project, analysed the data and co-wrote the paper. P.K.L conceived and designed the project, carried out the experiments, analysed the data and co-wrote the paper. P.K. and G.C made the confocal fluorescence and TIRF microscopy measurements and analysed the data. F.A.A. assisted in project design. C.K.M. carried out the cryo-EM experiment. G.D.H. helped with the design of nanotube 13 and with the graphical illustrations.

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The authors declare no competing financial interests.

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