www.sciencemag.org/cgi/content/full/science.aaf4388/DC1 Supplementary Materials for Designer nanoscale DNA assemblies programmed from the top down Rémi Veneziano, Sakul Ratanalert, Kaiming Zhang, Fei Zhang, Hao Yan, Wah Chiu, Mark Bathe* *Corresponding author. Email: [email protected]Published 26 May 2016 on Science First Release DOI: 10.1126/science.aaf4388 This PDF file includes: Materials and Methods Supplementary Text S1 to S7 Figs. S1 to S59 Tables S1 to S5 Captions for tables S6 to S27 Captions for movies S1 to S7 References (57–64) Other Supplementary Materials for this manuscript include the following: (available at www.sciencemag.org/cgi/content/full/science.aaf4388/DC1) Tables S6 to S27 Movies S1 to S7 DAEDALUS Software Package (zip file)
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Materials and Methods Supplementary Text S1 to S7 Figs. S1 to S59 Tables S1 to S5 Captions for tables S6 to S27 Captions for movies S1 to S7 References (57–64)
Other Supplementary Materials for this manuscript include the following: (available at www.sciencemag.org/cgi/content/full/science.aaf4388/DC1)
Tables S6 to S27 Movies S1 to S7 DAEDALUS Software Package (zip file)
2
Materials and Methods:
Design Algorithm
Specifying geometry
The goal of this work is to design and synthesize scaffolded DNA origami structures with
a top-down approach: given a target 3D structure of specified size and geometry (Fig. 1), the
algorithm will route a single-stranded scaffold throughout the entire geometry and generate the
required staple strands needed to experimentally fold the structure. We term our algorithm
DAEDALUS (DNA Origami Sequence Design Algorithm for User-defined Structures) (see
online Supporting Software and http://daedalus-dna-origami.org) for its ability to fully
automatically generate the scaffold routing and complementary staple sequences for an arbitrary
3D shape. 3D geometries are specified using a closed surface that is discretized using a
polyhedral mesh. In order to specify the geometry for scaffold routing, the spatial coordinates of
all vertices, the edge connectivities between vertices, and the faces to which vertices belong must
be provided (Fig. 2A and Fig. S1A to S1B). These may be provided manually, or through a file
format that specifies polygonal geometry, such as the Polygon File Format (PLY),
Stereolithography (STL), or Virtual Reality Modeling Language (WRL). As explained in more
detail below, any closed, orientable surface network can serve as input to the algorithm (Fig. 2
and Fig. S4 to S5). Provided in the code is a parser to convert PLY files into the required inputs.
In addition to the preceding spatial information, the desired minimum edge length, in bp, must be
specified for the structure (e.g. 31 or 42 bp). Each edge of the final scaffolded DNA origami
structure must be a multiple of 10.5 bp, rounded up or down to the nearest nucleotide, with a
minimum of 31 bp. For structures with equal edge lengths throughout the geometry, such as
Platonic, Archimedean, or Johnson solids, the desired minimum edge length simply becomes the
length for every edge. For other geometries, rounding edge lengths may be required, resulting in
some possible deviation between the specified target structure and final design. In these cases,
the desired minimum edge length is assigned to the shortest edge and the other edges are scaled
and rounded appropriately. When using the automated rounding to generate edge lengths, the
user is advised to verify that edge lengths are satisfactory before proceeding to the scaffold
routing procedure. If they are not, or if the user desires a structure with the same edge and face
connectivities but different edge lengths (e.g. modifying a regular tetrahedron into an irregular
tetrahedron), the user can edit and specify each edge length individually, so long as the lengths
Fig. S34. Characterization of the 52-bp edge-length DNA cube. The DNA cube was folded in
TAE-Mg2+
(12 mM MgCl2) buffer using the 1,616-nt scaffold amplified using aPCR and
characterized using agarose gel electrophoresis, AFM, and cryo-EM. Scale bars are 5 nm for the
model and 20 nm for AFM and cryo-EM.
46
Fig. S35. AFM imaging of 52-bp edge-length DNA cube. Image size 2 μm × 2 μm.
Fig. S36. Cryo-EM imaging of 52-bp edge-length DNA cube.
47
S4 Effects of salt on folding of DNA nanostructures
This section contains the full AFM images of the inset presented in Fig. 6 to 7 for the 52-bp edge
length pentagonal bipyramid DNA origami and additional results obtained with tetrahedron 63-
bp edge length. In order to completely characterize the folding of our DNA nanostructures, TAE
buffers containing different MgCl2 concentration (in a range of 0 to 30 mM) or NaCl (in a range
of 0 mM to 2 M) were used to fold tetrahedron 63-bp edge length or pentagonal bipyramid 52-bp
edge length. The characterization was realized using agarose gel electrophoresis and AFM
imaging.
Folding of pentagonal bipyramid in increasing MgCl2 concentration
Fig. S37. AFM imaging of 52-bp edge-length DNA pentagonal bipyramid folded in 1 mM
MgCl2. Image size 2 μm × 2 μm.
48
Fig. S38. AFM imaging of 52-bp edge-length DNA pentagonal bipyramid folded in 4 mM
MgCl2. Image size 2 μm × 2 μm.
Fig. S39. AFM imaging of 52-bp edge-length DNA pentagonal bipyramid folded in 30 mM
MgCl2. Image size 2 μm × 2 μm.
49
Folding of the DNA pentagonal bipyramid in increasing NaCl concentration
Fig. S40. AFM imaging of 52-bp edge-length DNA pentagonal bipyramid folded in 10 mM
NaCl. Image size 2 μm × 2 μm.
50
Fig. S41. AFM imaging of 52-bp edge-length DNA pentagonal bipyramid folded in 150 mM
NaCl. Image size 2 μm × 2 μm.
Fig. S42. AFM imaging of 52-bp edge-length DNA pentagonal bipyramid folded in 1 M
NaCl. Image size 2 μm × 2 μm.
51
Folding of the 63-bp edge-length tetrahedron in increasing concentration of MgCl2 and NaCl
Fig. S43. Folding of 63-bp edge-length DNA tetrahedron in buffer containing various
concentration of salt. The 63-bp edge-length DNA tetrahedron was folded in TRIS buffer with
increasing concentration of MgCl2 (0–30 mM) or increasing concentration of NaCl (0.01–2 M).
Folding was characterized with agarose gel electrophoresis and AFM imaging.
52
Fig. S44. AFM imaging of 63-bp edge-length DNA tetrahedron folded in 1 mM MgCl2. Image size 2 μm × 2 μm.
Fig. S45. AFM imaging of 63-bp edge-length DNA tetrahedron folded in 30 mM MgCl2. Image size 2 μm × 2 μm.
53
Fig. S46. AFM imaging of 63-bp edge-length DNA tetrahedron folded in 0.01 M NaCl. Image size 2 μm × 2 μm.
Fig. S47. AFM imaging of 63-bp edge-length DNA tetrahedron folded in 0.15 M NaCl.
Image size 2 μm × 2 μm.
54
Fig. S48. AFM imaging of 63-bp edge-length DNA tetrahedron folded in 1 M NaCl. Image
size 2 μm × 2 μm.
55
S5 Effects of buffer composition on folding of DNA origami objects
Fig. S49. Agarose gel of 52-bp edge-length DNA pentagonal bipyramid folded in PBS or
TAE buffer.
Fig. S50. AFM imaging of 52-bp edge-length DNA pentagonal bipyramid folded in PBS
buffer. Image size 2 μm × 2 μm.
56
Fig. S51. AFM imaging of 52-bp edge-length DNA pentagonal bipyramid folded in TAE
buffer. Image size 2 μm × 2 μm.
Fig. S52. AFM imaging of 52-bp edge-length DNA pentagonal bipyramid folded in TRIS-
NaCl 150 mM with phosphate (10 mM) and KCl (3 mM). Image size 2 μm × 2 μm.
57
Fig. S53. Agarose gel of 63-bp edge-length DNA tetrahedron folded in PBS or TAE buffer.
58
S6 Stability of DNA origami objects in physiological conditions
Fig. S54. AFM imaging of 52-bp edge-length DNA pentagonal bipyramid folded in TAE-
Mg2+
after buffer exchange with PBS. Image size 2 μm × 2 μm.
59
Fig. S55. AFM imaging of 52-bp edge-length DNA pentagonal bipyramid folded in TAE-
Mg2+
after buffer exchange with DMEM buffer. Image size 2 μm × 2 μm.
Fig. S56. AFM imaging of 52-bp edge-length DNA pentagonal bipyramid folded in TAE-
Mg2+
after buffer exchange with DMEM 2% dFBS. Image size 2 μm × 2 μm.
60
Fig. S57. AFM imaging of the 52-bp edge-length DNA pentagonal bipyramid folded in
TAE-Mg2+
after buffer exchange with DMEM 10% FBS. Image size 2 μm × 2 μm.
Fig. S58. Agarose gel electrophoresis of the 63-bp edge-length DNA tetrahedron to
determine its stability in DMEM with different concentrations of FBS for 6 hours.
61
Fig. S59. AFM imaging of 52-bp edge-length DNA pentagonal bipyramid folded in TAE-
Mg2+
after buffer exchange with TAE. Image size 2 μm × 2 μm.
62
Additional Tables (as a zipped archive):
Table S6. aPCR amplified sequences.
Table S7. Digested sequences.
Tables S8 to S27. Staple lists for the scaffolded DNA origami objects synthesized in this
paper.
Movies:
Movie S1. 3D rotation of icosahedron cryo-EM map with atomic model superimposed.
Movie S2. 3D rotation of tetrahedron cryo-EM map with atomic model superimposed.
Movie S3. 3D rotation of cuboctahedron cryo-EM map with atomic model superimposed.
Movie S4. 3D rotation of octahedron cryo-EM map with atomic model superimposed.
Movie S5. 3D rotation of reinforced cube cryo-EM map with atomic model superimposed.
Movie S6. 3D rotation of nested cube cryo-EM map with atomic model superimposed.
Movie S7. 3D rotation of octahedron cryo-EM map with alternate atomic model
superimposed. Before map-fitting. See Fig. S5(B), left.
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