www.sciencemag.org/cgi/content/full/338/6106/506/DC1

Supplementary Materials for

Fluorescence Enhancement at Docking Sites of DNA-Directed Self-Assembled Nanoantennas

G. P. Acuna,* F. M. Möller, P. Holzmeister, S. Beater, B. Lalkens, P. Tinnefeld*

*To whom correspondence should be addressed. E-mail: g.acuna@tu-bs.de (G.P.A.); p.tinnefeld@tu-bs.de (P.T.)

Published 26 October 2012, Science 338, 506 (2012) DOI: 10.1126/science.1228638

This PDF file includes:

Materials and Methods Figs. S1 to S9 Table S1 Full Reference List

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Materials and Methods Numerical simulations:

Numerical simulations were performed using commercial FDFD software (www.cst.com). The relative change in the excitation rate was estimated by simulating the electric field intensity in the vicinity of the nanoparticle system at the fluorophore position and averaged over the tangential and radial incident polarization. The relative change of the radiative and non radiative decay as well as the relative change in the lifetime was estimated following ref. (30), where the dye is modeled by a dipole current source with a determined orientation. The total power radiated into the far field and dissipated by the metallic objects is computed and normalized to the total power radiated into the far field in the absence of metallic objects. For these calculations, the intrinsic quantum yield of the ATTO647N dye of 0.65 was considered according to (31). All simulations were performed assuming a planar illumination with water as medium, mimicking the buffer conditions and effects arising from the glass interface were neglected. The error bars take into account the size distribution of the nanoparticles, but the deviation of particles from the spherical shape, the angular distribution of the DNA origami pillar or the possibility that not all three capturing strands are binding to a nanoparticle are not considered.

Functionalization of gold nanoparticles:

Gold nanoparticles of five different diameters (20, 40, 60, 80 and 100 nm) were purchased from BBInternational (www.bbi-gold.com). TEM measurements revealed the following size distribution, (19.4 ± 1.7 nm, 43.5 ± 4.6 nm, 61.5 ± 7 nm, 80.1 ± 7.4 nm, 105 ± 10.4 nm). Additionally some particles showed a considerable deviation from a spherical shape, see Fig. S3.

The DNA functionalization was performed as described in ref (32) with the following DNA sequence from VBC Biotech (www.vbc-biotech.com) containing a thiol modification at the 5’ end: 5’-Thiol-TTTTTTTTTTTTTTT-3’. The binding of the gold nanoparticles to the DNA origami structure was included in ref (19). The immobilized DNA origami pillars were incubated with DNA modified gold nanoparticles until the desired binding yield was achieved. This incubation was usually carried out on the microscope for several hours. Determination of intensity and fluorescence lifetime:

Fluorescence intensity and lifetime measurements were carried out on a custom-built confocal setup based on an Olympus IX-71 inverted microscope. For excitation, the light of an 80 MHz pulsed diode laser (640nm, LDH-D-C-640, Picoquant) was coupled into an oil-immersion objective (UPlanSApo 60XO / 1.35 NA, Olympus). A linear polarizer, an electrooptical modulator (EOM, LM 0202, Qioptiq) and a quarter wave plate (AQWP05M-600, Thorlabs) in the correct orientation with respect to each other allowed control over the excitation polarization (33). Depending on the rotation speed, we either measured with rotating linear polarization or average over all directions which for our measurements is equivalent to measurements with circular polarized light. Excitation and emission light were separated by a dual-band dichroic beam splitter (z532/633, AHF) and subsequently focused onto a 50 µm pinhole (Linos) before detection with an avalanche

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photo diode (τ-SPAD-100, Picoquant) with appropriate spectral filtering (ET 700/75m, AHF and RazorEdge LP 647, Semrock). The detector signal was registered with a single photon counting PC card (SPC-830, Becker&Hickl) and further analyzed using custom-made LabView code.

For FRET measurements, an additional 532 nm laser (TECGL-30, World Star Tech) was used to excite Cy3 and the emitted light was split spectrally at 640 nm (640DCXR, AHF) after the pinhole onto a second detector (τ-SPAD-100, Picoquant) with appropriate filters (Brightline HC582/75, AHF and RazorEdge LP 532, Semrock). The two lasers were alternated with 1 ms period by use of an acousto-optical tunable filter (AOTFnc-VIS, AA optoelectronic) to separate FRET sensitized from direct Cy5 excitation.

We employed the reconvolution algorithm of the FluoFit software (Picoquant) to obtain the fluorescence lifetime from the measured decay and the instrument response function (IRF) of the setup. We used an IRF acquired at an appropriate intensity to account for count rate dependence of the detector and included the periodicity of the excitation as well as scattering in the analysis. The width of the IRF (FWHM=650 ps) limits the temporal resolution to approximately 100-200 ps. All decays could be fitted with the convolution of a monoexponential decay and the IRF.

The Holliday junction:

The Holliday junction consisted of four strands (28) called R, H, X and B with the following sequences R: 5’- ACA AAT ATC CTT GCC CCA GCA GGC GAA TTT CCC ACC GCT CGG CTC AAC TGG G -3’, H: 5’-Cy3-CCG TAG CAG CGCG AGC GGT GGG-3’, X: 5’-CCC AGT TGA GCG CTT GCT AGG G-3’ and B: 5’-Cy5-CCC TAG CAA GCC GCT GCT AGG G-3’. The first 27 base pairs of the strand R bind to the origami pillar. All four strands were incorporated together with the staples prior to the folding process.

DNA origami pillar structures:

Unmodified and modified staple strands (see Table S1) were purchased from MWG (Munich, Germany) or IBA (Göttingen, Germany) at a concentration of 100 μM and were used without further purification. DNA origamis are formed with a molar ratio of 1:30 between the viral DNA and the unmodified staple strands and 1:100 between the viral DNA and the modified staple strands. For preparation of the scaffold strands Escherichia coli strain K91 was infected with the respective M13mp18 phage (p8634) at a Multiplicity of Infection of ~1. After amplification, the phage particles were separated, purified and their ssDNA was extracted and purified similar as described before (17). The concentration was adjusted to 100 nM using a molecular weight of 330 g/mol per base and an extinction coefficient = 33 mg/ml for A260 = 1 in a NanoDrop Spectrophotometer (Peqlab, Erlangen, Germany). The DNA origami design was performed with the open-source software caDNAno (www.cadnano.org)(17). The folding buffer contained 12.5 mM MgCl2 as well as 5 mM Tris + 1 mM EDTA (pH 7.9 at 20°C). A TEM image of the DNA origami pillar is included in Fig. S8. Folding time was three days.

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Fig. S1. Sketch of the top-view of the DNA origami pillar. The numbered circles represent DNA helices. Helices 0 to 11 form the central 12-helix bundle. The remaining helixes form the extra 6-helix bundles of the base. The helix center-to-center distance is 3 nm (16). The positions of the single dye (ATTO647N), the capturing paint strands, the Holliday Junction and the nanoparticles (NP) are also included.

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Fig. S2. Fluorescence lifetime imaging microscopy of DNA origami pillars with 80 nm nanoparticles. Orange-red spots represent ATTO647N dyes on DNA origami pillars without nanoparticles. The shortened fluorescence lifetime (blue spots) indicates binding of nanoparticles. Commonly, intermediate binding yields were intended to use the fluorescence from DNA origami pillars without nanoparticles as internal reference (A). On the other hand, yields exceeding 70% for the dimer could be reached even for 80 nm particles (B). (C) Histogram of yields of dimers, monomers and DNA origamis without a nanoparticle for the measurement included in Fig. 3A.

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Fig. S3. (A)-(E) TEM images of the gold nanoparticles of five different sizes. Analysis of the images revealed some deviation from perfect spherical shapes as well as a distribution of sizes (19.4 ± 1.7 nm, 43.5 ± 4.6 nm, 61.5 ± 7 nm, 80.1 ± 7.4 nm, 105 ± 10.4 nm).

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Fig. S4. (A)-(E) Fluorescence intensity versus lifetime plot of the DNA origami pillar with binding sites for one (monomer) and two (dimer) particles of different diameters.

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Fig. S5. Simulated quantum yield (A) and fluorescence lifetime (B) as a function of the nanoparticle diameter for the monomer and dimer system at different dye orientations, radial and tangential. The error bars take into account the size distribution of the nanoparticles. The quantum yield and fluorescence lifetime is normalized to the properties of the dye in the absence of nanoparticles. A quantum yield of 0.65 for ATTO647N was considered for both the quantum yield and fluorescence lifetime calculations.

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Fig. S6. Time gating for signal to noise improvement at elevated background fluorescence from freely diffusing dye molecules. (A) Fluorescence decay of the ATTO647N dye located within the DNA origami pillar with two 80 nm nanoparticles (dimer). Measurements were carried out at a concentration of 0.5 µM of ATTO647N dyes in solution. The decay shows a bi-exponential behavior due to the combination of the quenched dye in the hotspot and the unquenched dyes of the background. (B) Corresponding intensity transient showing that the single ATTO647N in the hotspot is easily detected despite the high background concentration. For (C) and (D) the intensity transients were reconstructed by time gating allowing only photons arriving 1.5-6 ns and 1.5-3 ns in the fluorescence decay, respectively. The time gating further increased the signal-to-noise by removing the largest fraction of the background fluorescence.

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Fig. S7. High count rates for fast dynamics. (A) Intensity transient at 1 µs binning. Off-states as short as 10 µs are clearly visualized. Therefore, the excitation intensity was increased to 7 µW and 100 µM ascorbic acid and 100 µM of methylviologen were added to the buffer to induce fast blinking of the dye molecule (34). At these high count rates, saturation effects of the setup and not by the organic dye limit photon counts. To estimate the actual rate of photons arriving at the detector in the on-state, we corrected for the dead time of the setup (B,C). Therefore, we measured the detected count rate of a micromolar concentration of ATTO647N in solution for different excitation intensities in an excitation regime where fluorescence from ATTO647N is linearly depending on excitation intensity (B). Photon count rate vs excitation intensity for a µM concentration of ATTO647N. (C) Correction factor dependence on the measured count rate. Hyperbolic fitting yields a saturation at Nmax = 6.8 ± 0.2 MHz which corresponds to a dead time of τD = 1/Nmax = 147 ± 4 ns. This yields a correction factor of f=1/(1-N τD) as displayed in (C). Accordingly, the count rate of the on-state for the transient displayed in (A) is 10.6 MHz. It should be emphasized that the blinking of the dye was purposely induced for this measurement and that all measurements discussed before were taken at lower excitation intensities to avoid saturation effects.

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Fig. S8. Transmission electron microscopy image of the DNA origami pillar.

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TATGCCGACTCTATATCTATACCTTCAT

CGTTCGGTATTTTTAATGGCGATGTTTTAGGGCTATCAGTTCGCGCATTAAAGACTAATAGCCATTCAAAAATATTGTCTGTGCCACGTATTCTTACGCTTTCAGGTCAGAAGGGTTCTATCTCTGTTGGCCAGAATGTCCCTTTTATTACTGGTCGTGTGACTGGTGAATCTGCCAATGTAAATAATCCATTTCAGACGATTGAGCGTCAAAATGTAGGTATTTCCATGAGCGTTTTTCCTGTTGCAATGGCTGGCGGTAATATTGTTCTGGATATTACCAGCAAGGCCGATAGTTTGAGTTCTTCTACTCAGGCAAGTGATGTTATTACTAATCAAAGAAGTATTGCTACAACGGTTAATTTGCGTGATGGACAGACTCTTTTACTCGGTGGCCTCACTGATTATAAAAACACTTCTCAGGATTCTGGCGTACCGTTCCTGTCTAAAATCCCTTTAATCGGCCTCCTGTTTAGCTCCCGCTCTGATTCTAACGAGGAAAGCACGTTATACGTGCTCGTCAAAGCAACCATAGTACGCGCCCTGTAGCGGCGCATTAAGCGCGGCGGGTGTGGTGGTTACGCGCAGCGTGACCGCTACACTTGCCAGCGCC

TTATAAACCTTCATGGAATATTTG

TGATAGACGGTTTTTCGCCCTTTGACGTTGGAGTCCACGTTCTTTAATAGTGGACTCTTGTTCCAAACTGGAACAACACTCAACCCTATCTCGGGCTATTCTTTTGATTTATAAGGGATTTTGCCGATTTCGGAACCACCATCAAACAGGATTTTCGCCTGCTGGGGCAAACCAGCGTGGACCGCTTGCTGCAACTCTCTCAGGGCCAGGCGGTGAAGGGCAATCAGCTGTTGCCCGTCTCACTGGTGAAAAGAAAAACCACCCTGGCGCCCAATACGCAAACCGCCTCTCCCCGCGCGTTGGCCGATTCATTAATGCAGCTGGCACGACAGGTTTCCCGACTGGAAAGCGGGCAGTGAGCGCAACGCAATTAATGTGAGTTAGCTCACTCATTAGGCACCCCAGGCTTTACACTTTATGCTTCCGGCTCGTATGTTGTGTGGAA

CTAGCGCCCGCTCCTTTCGCTTTCTTCCCTTCCTTTCTCGCCACGTTCGCCGGCTTTCCCCGTCAAGCTCTAAATCGGGGGCTCCCTTTAGGGTTCCGATTTAGTGCTTTACGGCACCTCGACCCCAAAAAACTTGATTTGGGTGATGGTTCACGTAGTGGGCCATCGCCC

GCCTAAATTACATGTTGGCGTTGTTAAATATGGCGATTCTCAATTAAGCCCTACTGTTGAGCGTTGGCTTTATACTGGTAAGAATTTGTATAACGCATATGATACTAAACAGGCTTTTTCTAGTAATTATGATTCCGGTGTTTATTCTTATTTAACGCCTTATTTATCACACGGTCGGTATTTCAAACCATTAAATTTAGGTCAGAAGATGAAATTAACTAAAATATATTTGAAAAAGTTTTCTCGCGTTCTTTGTCTTGCGATTGGATTTGCATCAGCATTTACATATAGTTATATAACCCAACCTAAGCCGGAGGTTAAAAAGGTAGTCTCTCAGACCTATGATTTTGATAAATTCACTATTGACTCTTCTCAGCGTCTTAATCTAAGCTATCGCTATGTTTTCAAGGATTCTAAGGGAAAATTAATTAATAGCGACGATTTACAGAAGCAAGGTTATTCACTCACATATATTGATTTATGTACTGTTTCCATTAAAAAAGGTAATTCAAATGAAATTGTTAAATGTAATTAATTTTGTTTTCTTGATGTTTGTTTCATCATCTTCTTTTGCTCAGGTAATTGAAATGAATAATTCGCCTCTGCGCGATTTTGTAACTTGGTATTCAAAGCAATCAGG

CTCTTATTACTGGCTCGAAAATGCCTCT

CTACATA

CGAATCCGTTATTGTTTCTCCCGATGTAAAAGGTACTGTTACTGTATATTCATCTGACGTTAAACCTGAAAATCTACGCAATTTCTTTATTTCTGTTTTACGTGCAAATAATTTTGATATGGTAGGTTCTAACCCTTCCATTATTCAGAAGTATAATCCAAACAATCAGGATTATATTGATGAATTGCCATCATCTGATAATCAGGAATATGATGATAATTCCGCTCCTTCTGGTGGTTTCTTTGTTCCGCAAAATGATAATGTTACTCAAACTTTTAAAATTAATAACGTTCGGGCAAAGGATTTAATACGAGTTGTCGAATTGTTTGTAAAGTCTAATACTTCTAAATCCTCAAATGTATTATCTATTGACGGCTCTAATCTATTAGTTGTTAGTGCTCCTAAAGATATTTTAGATAACCTTCCTCAATTCCTTTCAACTGTTGATTTGCCAACTGACCAGATATTGATTGAGGGTTTGATATTTGAGGTTCAGCAAGGTGATGCTTTAGATTTTTCATTTGCTGCTGGCTCTCAGCGTGGCACTGTTGCAGGCGGTGTTAATACTGACCGCCTCACCTCTGTTTTATCTTCTGCTGGTGGTT

TTACCTTTTGTCGGTACTTTATATT

AACACCTTCGTGAT

ACTCAAGGCACTGACCCCGTTAAAACTTATTACCAGTACACTCCTGTATCATCAAAAGCCATGTATGACGCTTACTGGAACGGTAAATTCAGAGACTGCGCTTTCCATTCTGGCTTTAATGAGGATTTATTTGTTTGTGAATATCAAGGCCAATCGTCTGACCTGCCTCAACCTCCTGTCAATGCTGGCGGCGGCTCTGGTGGTGGTTCTGGTGGCGGCTCTGAGGGTGGTGGCTCTGAGGGTGGCGGTTCTGAGGGTGGCGGCTCTGAGGGAGGCGGTTCCGGTGGTGGCTCTGGTTCCGGTGATTTTGATTATGAAAAGATGGCAAACGCTAATAAGGGGGCTATGACCGAAAATGCCGATGAAAACGCGCTACAGTCTGACGCTAAAGGCAAACTTGATTCTGTCGCTACTGATTACGGTGCTGCTATCGATGGTTTCATTGGTGACGTTTCCGGCCTTGCTAATGGTAATGGTGCTACTGGTGATTTTGCTGGCTCTAATTCCCAAATGGCTCAAGTCGGTGACGGTGATAATTCACCTTTAATGAATAATTTCCGTCAATATTTACCTTCCCTCCCTCAATCGGTTGAATGTCGCCC

TTGTCGTCGTCTGGACAGAATTACT

TTTTGTCTTTGGCGCTGGTAAACCATATGAATTTTCTATTGATTGTGACAAAATAAACTTATTCCGTGGTGTCTTTGCGTTTCTTTTATATGTTGCCACCTTTATGTATGTATTTTCTACGTTTGCTAACATACTGCGTAATAAGGAGTCTTAATCATGCCAGTTCTTTTGGGTATTCCGTTATTATTGCGTTTCCTCGGTTTCCTTCTGGTAACTTTGTTCGGCTATCTGCTTACTTTTCTTAAAAAGGGCTTCGGTAAGATAGCTATTGCTATTTCATTGTTTCTTGCTCTTATTATTGGGCTTAACTCAATTCTTGTGGGTTATCTCTCTGATATTAGCGCTCAATTACCCTCTGACTTTGTTCAGGGTGTTCAGTTAATTCTCCCGTCTAATGCGCTTCCCTGTTTTTATGTTATTCTCTCTGTAAAGGCTGCTATTTTCATTTTTGACGTTAAACAAAAAATCGTTTCTTATTTGGATTGGGATAAATAATATGGCTGTTTATTTTGTAACTGGCAAATTAGGCTCTGGAAAGACGCTCGTTAGCGTTGGTAAGATTCAGGATAAAATTGTAGCTGGGTGCAAAATAGCAACTAATCTTGATTTAAGGCTT

GTTGAAATTAAACCATCTCAAGCCCAATTTACTACTCGTTCTGGTGTTTCTCGTCAGGGCAAGCCTTATTCACTGAATGAGCAGCTTTGTTACGTTGATTTGGGTAATGAATATCCGGTTCTTGTCAAGATTACTCTTGATGAAGGTCAGCCAGCCTATGCGCCTGGTCTGTACACCGTTCATCTGTCCTCTTTCAAAGTTGGTCAGTTCGGTTCCCTTATGATTGACCGTCTGCGCCTCGTTCCGGCTAAGTAACATGGAGCAGGTCGCGGATTTCGACACAATTTATCAGGCGATGATACAAATCTCCGTTGTACTTTGTTTCGCGCTTGGTATAATCGCTGGGGGTCAAAGATGAGTGTTTTAGTGTATTCTTTTGCCTCTTTCGTTTTAGGTTGGTGCCTTCGTAGTGGCATTACGTATTTTACCCGTTTAATGGAAACTTCCTCATGAAAAAGTCTTTAGTCCTCAAAGCCTCTGTAGCCGTTGCTACCCTCGTTCCGATGCTGTCTTTCGCTGCTGAGGGTGACGATCCCGCAAAAGCGGCCTTTAACTCCCTGCAAGCCTCAGCGACCGAATATATCGGTTATGCGTGGGCGATGGTTGTTGTCATTGTCGGCGCAACTATCGGTATCAAGCT

TCGCATAAGGTAATTCACAATGATTAAA

GTCTGCA

GTTTAAGAAATTCACCTCGAAAGCAAGCTGATAAACCGATACAATTAAAGGCTCCTTTTGGAGCCTTTTTTTTGGAGATTTTCAACGTGAAAAAATTATTATTCGCAATTCCTTTAGTTGTTCCTTTCTATTCTCACTCCGCTGAAACTGTTGAAAGTTGTTTAGCAAAATCCCATACAGAAAATTCATTTACTAACGTCTGGAAAGACGACAAAACTTTAGATCGTTACGCTAACTATGAGGGCTGTCTGTGGAATGCTACAGGCGTTGTAGTTTGTACTGGTGACGAAACTCAGTGTTACGGTACATGGGTTCCTATTGGGCTTGCTATCCCTGAAAATGAGGGTGGTGGCTCTGAGGGTGGCGGTTCTGAGGGTGGCGGTTCTGAGGGTGGCGGTACTAAACCTCCTGAGTACGGTGATACACCTATTCCGGGCTATACTTATATCAACCCTCTCGACGGCACTTATCCGCCTGGTACTGAGCAAAACCCCGCTAATCCTAATCCTTCTCTTGAGGAGTCTCAGCCTCTTAATACTTTCATGTTTCAGAATAATAGGTTCCGAAATAGGCAGGGGGCATTAACTGTTTATACGGGCACTGTT

ACGTCCTGACTGGTATAATGAGCCAGTTCTTAAAA

TGGAGACAAGACAC

AAGCTGGCTGGAGTGCGATCTTCCTGAGGCCGATACTGTCGTCGTCCCCTCAAACTGGCAGATGCACGGTTACGATGCGCCCATCTACACCAACGTGACCTATCCCATTACGGTCAATCCGCCGTTTGTTCCCACGGAGAATCCGACGGGTTGTTACTCGCTCACATTTAATGTTGATGAAAGCTGGCTACAGGAAGGCCAGACGCGAATTATTTTTGATGGCGTTCCTATTGGTTAAAAAATGAGCTGATTTAACAAAAATTTAATGCGAATTTTAACAAAATATTAACGTTTACAATTTAAATATTTGCTTATACAATCTTCCTGTTTTTGGGGCTTTTCTGATTATCAACCGGGGTACATATGATTGACATGCTAGTTTTACGATTACCGTTCATCGATTCTCTTGTTTGCTCCAGACTCTCAGGCAATGACCTGATAGCCTTTGTAGATCTCTCAAAAATAGCTACCCTCTCCGGCATTAATTTATCAGCTAGAACGGTTGAATATCATATTGATGGTGATTTGACTGTCTCCGGCCTTTCTCACCCTTTTGAATCTTTACCTACACATTACTCAGGCATTGCATTTAAAA

TTAGTTCGTTTTATTAACGTAGATTTTTCTTCCCA

TATATGAGGGTTCTAAAAATTTTTATCCTTGCGTTGAAATAAAGGCTTCTCCCGCAAAAGTATTACAGGGTCATAATGTTTTTGGTACAACCGATTTAGCTTTATGCTCTGAGGCTTTATTGCTTAATTTTGCTAATTCTTTGCCTTGCCTGTATGATTTATTGGATGTTAATGCTACTACTATTAGTAGAATTGATGCCACCTTTTCAGCTCGCGCCCCAAATGAAAATATAGCTAAACAGGTTATTGACCATTTGCGAAATGTATCTAATGGTCAAACTAAATCTACTCGTTCGCAGAATTGGGAATCAACTGTTATATGGAATGAAACTTCCAGACACCGTACTTTAGTTGCATATTTAAAACATGTTGAGCTACAGCATTATATTCAGCAATTAAGCTCTAAGCCATCCGCAAAAATGACCTCTTATCAAAAGGAGCAATTAAAGGTACTCTCTAATCCTGACCTGTTGGAGTTTGCTTCCGGTCTGGTTCGCTTTGAAGCTCGAATTAAAACGCGATATTTGAAGTCTTTCGGGCTTCCTCTTAATCTTTTTGATGCAATCCGCTTTGCTTCTGACTATAATAGTCAGGGTAAAGACCTGATTT

TAACTCATTGATACTCATTTATAAACTCCTTGCAATGTATGTCGTTTCAGCTAAACGGTATCAGCAATGTTTATGTAAAGAAACAGTAAGATAATACTCAACCCGATGTTTGAGTACGGTCATCATCTGACACTACAGACTCTGGCATCGCTGTGAAGACGACGCGAAATTCAGCATTTTCACAAGCGTTATCTTTTACAAAACCGATCTCACTCTCCTTTGATGCGAATGCCAGCGTCAGACATCATATGCAGATACTCACCTGCATCCTGAACCCATTGACCTCCAACCCCGTAATAGCGATGCGTAATGATGTCGATAGTTACTAACGGGTCTTGTTCGATTAACTGCCGCAGAAACTCTTCCAGGTCACCAGTGCAGTGCTTGATAACAGGAGTCTTCCCAGGATGGCGAACAACAAGAAACTGGTTTCCGTCTTCACGGACTTCGTTGCTTTCCAGTTTAGCAATACGCTTACTCCCATCCGAGATAACACCTTCGTAATACTCACGCTGCTCGTTGAGTTTTGATTTTGCTGTTTCAAGCTCAACACGCAGTTTCCCTACTGTTAGCGCAATATCCTCGTTCTCCTGGTCGCGGCGTTTGATGTATTGCTGGTTTCTTTCCCGTTCATCCAGCAG

CGGATCTGCACAACATTGATAACGCCCAATCTTTTTGCTCAGACTC

TTCCAGCACAATCGATGGTGTTACCAATTCATGGAAAAGGTCTGCGTCAAATCCCCAGTCGTCATGCATTGCCTGCTCTGCCGCTTCACGCAGTGCCTGAGAGTTAATTTCGCTCACTTCGAACCTCTCTGTTTACTGATAAGTTCCAGATCCTCCTGGCAACTTGCACAAGTCCGACAACCCTGAACGACCAGGCGTCTTCGTTCATCTATCGGATCGCCACACTCACAACAATGAGTGGCAGATATAGCCTGGTGGTTCAGGCGGCGCATTTTTATTGCTGTGTTGCGCTGTAATTCTTCTATTTCTGATGCTGAATCAATGATGTCTGCCATCTTTCATTAATCCCTGAACTGTTGGTTAATACGCATGAGGGTGAATGCGAATAATAAAGCTTGGCACTGGCCGTCGTTTTACAACGTCGTGACTGGGAAAACCCTGGCGTTACCCAACTTAATCGCCTTGCAGCACATCCCCCTTTCGCCAGCTGGCGTAATAGCGAAGAGGCCCGCACCGATCGCCCTTCCCAACAGTTGCGCAGCCTGAATGGCGAATGGCGCTTTGCCTGGTTTCCGGCACCAGAAGCGGTGCCGGA

TGATGAATCTTTCTACCTGTAATAATGTTGTTCCG

TTGATTTATGGTCATTC

TGCATTAGTTGAATGTGGTATTCCTAAATCTCAAC

TCGTTTTCTGAACTGTT

CTCGTAATTCCTTTTGGCGTTATGTATC

TAAAGCATTTGAGGGGGATTCAAT

ATGATAGTGTTGCTCTTACTATGC

GAATATTTATGACGATTCCGCAGTATTG

TTTTATCGTCGTCTGGTAAACGAGGGTT

GACGCTATCCAGTCTAAACATTTTA

CTATTACCCCCTCTGGCAAAACTTCTTTTGCAAAAGCCTCTCGCTATTTTGGT

AATGGGGATTTGTCTTCATTAGAC

TTGTGAGCGGATAACAATTTCACACAGG

TCAGTCATTTCTCGCACATTGCAG

AAACAGCTATGACCATGATTACGAATTC

GTACTCGTGATAATAATTTTGCACGGTA

GAGCTCGGTACCCGGGGATCCATTC

AGAGAGGGCAAGTATCGTTTCCACC

TCCTGTGACTCGGAAGTGCATTTATCAT

TGGGTTTAATCATCTATATGTTTTGTAC

CTCCATAAAACAAAACCCGCCGTA

GCGAGTTCAGATAAAATAAATCCCCGCGAGTGCGAGGATTGTTATGTAATAT

CTGCATTAGCTGAACATGTTGTTTA

CAAAACCTCCCGCAAGTCGGGAGGTTCG

TCTATTGTTGATAAACAGGCGCGTT

CTAAAACGCCTCGCGTTCTTAGAATACC

GGATATTATTTTTCTTGTTCAGGACTTA

GGATAAGCCTTCTATATCTGATTT

TCTACATGCTCGTAAATTAGGATG

GCTTGCTATTGGGCGCGGTAATGATTCC

CCGATTATTGATTGGTT

TACGATGAAAATAAAAACGGCTTGCTTGTTCTCGA

TGAGTGCGGTACTTGGTTTAATACCCGTTCTTGGAATGATAAGGAAAGACAG

TAGATATAGAGTCG

GAAGGTA

GAACGAT

AAATACC

CCATTAA

AACATCG

GCCCTAA

ACTGATA

TGCGCGA

TCTTTAA

CTATTAG

TGAATGG

ATATTTT

ACAGACA

ACGTGGC

TAAGAAT

GAAAGCG

CTGACCT

AACCCTT

GAGATAG

GCCAACA

CATTCTG

AAAGGGA

AGTAATA

CACGACC

CCAGTCA

AGATTCA

CATTGGC

TTATTTA

AAATGGA

TCGTCTG

CGCTCAA

ATTTTGA

TACCTAC

ATGGAAA

AACGCTC

CAGGAAA

ATTGCAA

GCCAGCC

TATTACC

AGAACAA

AATATCC

TGCTGGT

TCGGCCT

CAAACTA

AAGAACT

TGAGTAG

ACTTGCC

TAACATC

TTAGTAA

TCTTTGA

CAATACT

GTTGTAG

ATTAACC

CACGCAA

TGTCCAT

AAGAGTC

CGAGTAA

AGGCCAC

ATCAGTG

TTTTATA

GAAGTGT

ATCCTGA

CGCCAGA

AACGGTA

AGACAGG

GGATTTT

ATTAAAG

GAGGCCG

TAAACAG

CGGGAGC

ATCAGAG

CGTTAGA

CTTTCCT

TAACGTG

GCACGTA

TTGACGA

GGTTGCT

GTACTAT

AGGGCGC

CCGCTAC

TAATGCG

CCGCGCT

ACACCCG

AACCACC

TGCGCGT

GTCACGC

GGCAAGTGTAGCG

CAAATATTCCATGAAGGTTTATAATT

CCACACAACATACG

AGCCGGA

AGCATAAAGTGTAA

AGCCTGG

GGTGCCTAATGAGT

GAGCTAA

CTCACATTAATTGC

GTTGCGC

TCACTGCCCGCTTT

CCAGTCG

GGAAACCTGTCGTG

CCAGCTG

CATTAATGAATCGG

CCAACGC

GCGGGGAGAGGCGG

TTTGCGT

ATTGGGCGCCAGGG

TGGTTTT

TCTTTTCACCAGTG

AGACGGG

CAACAGCTGATTGC

CCTTCAC

CGCCTGGCCCTGAG

AGAGTTG

CAGCAAGCGGTCCA

CGCTGGT

TTGCCCCAGCAGGC

GAAAATC

CTGTTTGATGGTGG

TTCCGAA

ATCGGCAAAATCCC

TTATAAA

TCAAAAGAATAGCC

CGAGATA

GGGTTGAGTGTTGT

TCCAGTT

TGGAACAAGAGTCC

ACTATTA

AAGAACGTGGACTC

CAACGTC

AAAGGGCGAAAAAC

CGTCTAT

CAGGGCGATGGCCC

ACTACGT

GAACCATCACCCAA

ATCAAGT

TTTTTGGGGTCGAG

GTGCCGT

AAAGCACTAAATCG

GAACCCT

AAAGGGAGCCCCCG

ATTTAGA

GCTTGACGGGGAAA

GCCGGCG

AACGTGGCGAGAAA

GGAAGGG

AAGAAAGCGAAAGGAGCGGG

TAACAACGCCAACATGTAA

CCATATT

AGAATCG

TTAATTG

CTCAACAGTAGGGC

GCCAACG

CTTACCAGTATAAA

ACAAATT

ATCATATGCGTTAT

GTTTAGT

ACTAGAAAAAGCCT

CATAATT

ATAAACACCGGAAT

AATAAGA

TAAATAAGGCGTTA

CGTGTGA

TTTGAAATACCGAC

TTAATGG

CTTCTGACCTAAAT

ATTTCAT

ATATATTTTAGTTA

TTTTCAA

AACGCGAGAAAACT

GACAAAG

AAATCCAATCGCAA

CTGATGC

ACTATATGTAAATG

TTATATA

CGGCTTAGGTTGGG

TAACCTC

AGAGACTACCTTTT

AGGTCTG

TTTATCAAAATCAT

TAGTGAA

CTGAGAAGAGTCAA

TAAGACG

GCGATAGCTTAGAT

AAACATA

CTTAGAATCCTTGA

ATTTTCC

GTCGCTATTAATTA

GTAAATC

ATAACCTTGCTTCT

TGAGTGA

TAAATCAATATATG

CAGTACA

TTTTTTAATGGAAA

AATTACC

AACAATTTCATTTG

TACATTT

AAAACAAAATTAAT

CATCAAG

GATGATGAAACAAA

AAAAGAA

TCAATTACCTGAGC

ATTCATT

CGCAGAGGCGAATT

AAAATCG

TTGCTTTGAATACCAAGTTAC

AGAGGCATTTTCGAGCCAGTAATAAG

AGTATGTAGAACCACCAGCAG

AAGATAA

AACAGAG

GTGAGGC

GGTCAGT

ATTAACA

CCGCCTG

CAACAGT

GCCACGC

TGAGAGC

CAGCAGC

AAATGAA

AAATCTA

AAGCATC

ACCTTGC

TGAACCT

CAAATAT

CAAACCC

TCAATCA

ATATCTG

GTCAGTT

GGCAAAT

CAACAGT

TGAAAGG

AATTGAG

GAAGGTT

ATCTAAA

ATATCTT

TAGGAGC

ACTAACA

ACTAATA

GATTAGA

GCCGTCA

ATAGATA

ATACATT

TGAGGAT

TTAGAAG

TATTAGA

CTTTACA

AACAATT

CGACAAC

TCGTATT

AAATCCT

TTGCCCG

AACGTTA

TTAATTT

TAAAAGT

TTGAGTA

ACATTAT

CATTTTG

CGGAACA

AAGAAAC

CACCAGA

AGGAGCG

GAATTAT

CATCATA

TTCCTGA

TTATCAG

ATGATGG

CAATTCA

TCAATAT

AATCCTG

ATTGTTT

GGATTAT

ACTTCTG

AATAATG

GAAGGGT

TAGAACC

TACCATA

TCAAAAT

TATTTGC

ACGTAAA

ACAGAAA

TAAAGAA

ATTGCGT

AGATTTT

CAGGTTT

AACGTCA

GATGAAT

ATACAGT

AACAGTA

CCTTTTA

CATCGGG

AGAAACA

ATAACGG

TATAAAGTACCGACA

CACGAAGGTGTTAA

TGAGTAT

AGTGCCT

CGGGGTC

GTTTTAA

GTAATAA

TGTACTG

ACAGGAG

TGATGAT

TGGCTTT

TCATACA

GTAAGCG

CGTTCCA

AATTTAC

GTCTCTG

AAGCGCA

GAATGGA

AAAGCCA

CCTCATT

AATAAAT

ACAAACA

GATATTC

TGGCCTT

AGACGAT

GCAGGTC

GGTTGAG

GACAGGA

CAGCATT

CCGCCGC

ACCAGAG

AACCACC

CCACCAG

AGAGCCG

CACCCTC

GAGCCAC

ACCCTCA

AACCGCC

CCCTCAG

GCCGCCA

CCTCAGA

CCGCCTC

ACCGGAA

AGCCACC

GAACCAG

ATCACCG

AATCAAA

TTTTCAT

TGCCATC

TAGCGTT

CCCTTAT

CATAGCC

TTTCGGT

TCGGCAT

GTTTTCA

GTAGCGC

TCAGACT

TTTAGCG

GTTTGCC

GAATCAA

AGCGACA

AATCAGT

GCACCGT

GATAGCA

AACCATC

CCAATGA

AACGTCA

GGCCGGA

TTAGCAA

ATTACCA

TAGCACC

TCACCAG

AGCAAAA

TAGAGCC

TGGGAAT

AGCCATT

CGACTTG

CCGTCAC

ATTATCA

AAGGTGA

TTCATTA

GAAATTA

ATTGACG

GGTAAAT

GAGGGAA

ATTGAGG

ACATTCAACCG

AGTAATTCTGTCCAGACGACGACAAAAGCC

TTAAATC

AAGATTAGTTGCTA

TTTTGCA

CCCAGCTACAATTT

TATCCTG

AATCTTACCAACGC

TAACGAG

CGTCTTTCCAGAGC

CTAATTT

GCCAGTTACAAAAT

AAACAGC

CATATTATTTATCC

CAATCCA

AATAAGAAACGATT

TTTTGTT

TAACGTCAAAAATG

AAAATAG

CAGCCTTTACAGAG

AGAATAA

CATAAAAACAGGGA

AGCGCAT

TAGACGGGAGAATT

AACTGAA

CACCCTGAACAAAG

TCAGAGG

GTAATTGAGCGCTA

ATATCAG

AGAGATAACCCACA

AGAATTG

AGTTAAGCCCAATA

ATAAGAG

CAAGAAACAATGAA

ATAGCAA

TAGCTATCTTACCG

AAGCCCT

TTTTAAGAAAAGTA

AGCAGAT

AGCCGAACAAAGTT

ACCAGAA

GGAAACCGAGGAAA

CGCAATA

ATAACGGAATACCC

AAAAGAA

CTGGCATGATTAAG

ACTCCTT

ATTACGCAGTATGT

TAGCAAA

CGTAGAAAATACAT

ACATAAA

GGTGGCAACATATA

AAAGAAA

CGCAAAGACACCAC

GGAATAA

GTTTATTTTGTCAC

AATCAAT

AGAAAATTCATATG

GTTTACC

AGCGCCAAAGAC

TTTAATT

GAGATGG

TGGGCTT

AGTAAAT

ACACCAGAACGAGT

TGCCCTGACGAGAA

TAAGGCT

GCTCATTCAGTGAA

CAAAGCT

CCAAATCAACGTAA

TCATTAC

CAAGAACCGGATAT

ATCTTGA

CTTCATCAAGAGTA

GGCTGAC

CAGGCGCATAGGCT

TACAGAC

ACAGATGAACGGTG

AAAGAGG

ACTGACCAACTTTG

GAACCGA

GGTCAATCATAAGG

CGCAGAC

TAGCCGGAACGAGG

TGTTACT

CGCGACCTGCTCCA

CGAAATC

CTGATAAATTGTGT

TCATCGC

AACGGAGATTTGTA

AAAGTAC

ACCAAGCGCGAAAC

CGATTAT

TCTTTGACCCCCAG

ACACTCA

AAGAATACACTAAA

GAGGCAA

AACCTAAAACGAAA

AGGCACC

TAATGCCACTACGA

AAATACG

CCATTAAACGGGTA

GAAGTTT

GACTTTTTCATGAG

GACTAAA

ACAGAGGCTTTGAG

AACGGCT

GGAACGAGGGTAGC

CAGCATC

TCAGCAGCGAAAGA

GTCACCC

GCTTTTGCGGGATC

AAAGGCC

GCTTGCAGGGAGTT

CGCTGAG

CCGATATATTCGGT

CGCATAA

ACAACCATCGCCCA

AATGACA

ATACCGATAGTTGCGCCGAC

TTTAATCATTGT

GAATTAC

CTTATGC

GATGCAGACAACAG

TGCCCGT

ATAAACA

GTTAATG

CCCCCTG

CCTATTT

CGGAACC

TATTATT

CTGAAAC

ATGAAAG

TATTAAG

AGGCTGA

GACTCCT

CAAGAGA

AGGATTA

GGATTAG

CGGGGTT

TTGCTCA

GTACCAG

GCGGATA

AGTGCCG

TCGAGAG

GGTTGAT

ATAAGTA

TAGCCCG

GAATAGG

TGTATCA

CCGTACT

CAGGAGG

TTTAGTA

CCGCCAC

CCTCAGA

ACCGCCA

CCCTCAG

AACCGCC

ACCCTCA

GAGCCAC

CACCCTC

ATTTTCA

GGGATAG

CAAGCCC

AATAGGA

ACCCATG

TACCGTA

ACACTGA

GTTTCGT

CACCAGT

ACAAACT

ACAACGC

CTGTAGC

ATTCCAC

AGACAGC

CCTCATA

GTTAGCG

TAACGAT

CTAAAGT

TTTGTCG

TCTTTCC

AGACGTT

AGTAAAT

GAATTTT

CTGTATG

GGATTTT

GCTAAAC

AACTTTC

AACAGTT

TCAGCGG

AGTGAGA

ATAGAAA

GGAACAA

CTAAAGG

AATTGCG

AATAATA

ATTTTTT

CACGTTG

AAAATCT

CCAAAAA

AAAGGCT

CCAAAAG

GAGCCTT

TAATTGT

ATCGGTT

TATCAGC

TTGCTTT

CGAGGTGAATTTC

ATTATACCAGTCA

CTGGCTC

TTAAGAA

GTCTTGTCTCCATT

TCCAGCCAGCTTGT

ATCGCAC

CAGGAAG

TCGGCCT

GACAGTA

GGACGAC

TTTGAGG

CTGCCAG

CGTGCAT

TCGTAAC

GGGCGCA

TGTAGAT

ACGTTGG

ATAGGTC

TAATGGG

TTGACCG

CGGCGGA

GAACAAA

TCCGTGG

CGGATTC

AACCCGT

GAGTAAC

TGTGAGC

CATTAAA

TCATCAA

CCAGCTT

CCTGTAG

TGGCCTT

TCGCGTC

AAATAAT

CCATCAA

AGGAACG

AACCAAT

ATTTTTT

TCAGCTC

TGTTAAA

AAATTTT

TCGCATT

TTAAAAT

TATTTTG

ACGTTAA

ATTGTAA

TATTTAA

AAGCAAA

ATTGTAT

CAGGAAG

CCAAAAA

AAAAGCC

TAATCAG

CGGTTGA

TGTACCC

AATCATA

GCATGTC

AAAACTA

TAATCGT

TGAACGG

GAATCGA

AACAAGA

TGGAGCA

GAGAGTC

ATTGCCT

TCAGGTC

AAGGCTA

ATCTACA

TTGAGAG

GCTATTT

GAGGGTA

TGCCGGA

AAATTAA

AGCTGAT

CCGTTCT

TATTCAA

AATATGA

CACCATC

GTCAAAT

GGAGACA

AAAGGCC

GGGTGAG

TTCAAAA

GTAAAGA

TGTGTAG

TGAGTAA

CAATGCC

TGGGAAGAAAAATCTACGT

TAATAAAACGAACT

AAAAATCAGGTCTT

TACCCTGACTATTA

TAGTCAG

AAGCAAAGCGGATT

GCATCAA

AAAGATTAAGAGGA

AGCCCGA

AAGACTTCAAATAT

CGCGTTT

TAATTCGAGCTTCA

AAGCGAA

CCAGACCGGAAGCA

AACTCCA

ACAGGTCAGGATTA

GAGAGTA

CCTTTAATTGCTCC

TTTTGAT

AAGAGGTCATTTTT

GCGGATG

GCTTAGAGCTTAAT

TGCTGAA

TATAATGCTGTAGC

TCAACAT

GTTTTAAATATGCA

ACTAAAG

TACGGTGTCTGGAA

GTTTCAT

TCCATATAACAGTT

GATTCCC

AATTCTGCGAACGA

GTAGATT

TAGTTTGACCATTA

GATACAT

TTCGCAAATGGTCA

ATAACCT

GTTTAGCTATATTT

TCATTTG

GGGCGCGAGCTGAA

AAGGTGG

CATCAATTCTACTA

ATAGTAG

TAGCATTAACATCC

AATAAAT

CATACAGGCAAGGC

AAAGAAT

TAGCAAAATTAAGC

AATAAAG

CCTCAGAGCATAAA

GCTAAAT

CGGTTGTACCAAAA

ACATTAT

GACCCTGTAATACT

TTTGCGG

GAGAAGCCTTTATT

TCAACGC

AAGGATAAAAATTTTTAGAA

GAGTATCAAT

TATAAAT

AGGAGTT

CATTGCA

CGACATA

CCGTTTAGCTGAAA

AAACATTGCTGATA

TTTACAT

TATCTTACTGTTTC

TGAGTAT

CTCAAACATCGGGT

GACCGTA

TAGTGTCAGATGAT

GAGTCTG

TCACAGCGATGCCA

GTCGTCT

ATGCTGAATTTCGC

TGTGAAA

TAAAAGATAACGCT

GGTTTTG

AGGAGAGTGAGATC

GCATCAA

TGACGCTGGCATTC

TGATGTC

TGAGTATCTGCATA

ATGCAGG

TCAATGGGTTCAGG

TTGGAGG

TCGCTATTACGGGG

TTACGCA

AACTATCGACATCA

CGTTAGT

AATCGAACAAGACC

GGCAGTT

GGAAGAGTTTCTGC

GTGACCT

CAAGCACTGCACTG

CTGTTAT

TCCTGGGAAGACTC

TTCGCCA

CCAGTTTCTTGTTG

ACGGAAA

CGAAGTCCGTGAAG

AAAGCAA

TATTGCTAAACTGG

GTAAGCG

TATCTCGGATGGGA

AAGGTGT

GCGTGAGTATTACG

ACGAGCA

AAAATCAAAACTCA

AAACAGC

GCGTGTTGAGCTTG

GGAAACT

TGCGCTAACAGTAG

AGGATAT

CGACCAGGAGAACG

AACGCCG

CCAGCAATACATCA

AAAGAAA

GATGAACGGG

GAGTCTGAGCAAAAAG

ATTGGGC

GTTATCA

ATGTTGT

GCAGATC

CGTCCGG

CACCGCT

TCTGGTG

CCGGAAA

CCAGGCA

AAGCGCC

ATTCGCC

ATTCAGG

CTGCGCA

ACTGTTG

GGAAGGG

CGATCGG

TGCGGGC

CTCTTCG

CTATTAC

GCCAGCT

GGCGAAA

GGGGGAT

GTGCTGC

AAGGCGA

TTAAGTT

GGGTAAC

GCCAGGG

TTTTCCC

AGTCACG

ACGTTGT

AAAACGA

CGGCCAG

TGCCAAG

CTTTATT

ATTCGCA

TTCACCC

TCATGCG

TATTAAC

CAACAGT

TCAGGGA

TTAATGA

AAGATGG

CAGACAT

CATTGAT

TCAGCAT

CAGAAAT

AGAAGAA

TTACAGC

GCAACAC

AGCAATA

AAAATGC

GCCGCCT

GAACCAC

CAGGCTA

TATCTGC

CACTCAT

TGTTGTG

AGTGTGG

CGATCCG

ATAGATG

AACGAAG

ACGCCTG

GTCGTTC

AGGGTTG

TCGGACT

TGTGCAA

GTTGCCA

GGAGGAT

CTGGAAC

TTATCAG

TAAACAG

AGAGGTT

CGAAGTG

AGCGAAA

TTAACTC

TCAGGCA

CTGCGTG

AAGCGGC

AGAGCAG

GCAATGC

ATGACGA

CTGGGGA

TTTGACG

CAGACCT

TTTCCAT

GAATTGG

TAACACC

ATCGATTGTG

CAGGTAGAAAGAT

ATTATTA

GAACAAC

ACCATAAATCAACG

GTTGAGATTTAGGAATACCACATTCA

ACTAATGCAAACAGTTCAGAA

TACATAACGCCAAAAGGAAT

ATCCCCCTCAAATGCTTTAGA

GCATAGTAAGAGCAACACTATCATCA

ATACTGCGGAATCGTCATAAA

CCAGACGACG

TCGTTTA

GATAGCGTCAACCC

ATGTTTAGACTG

ACCAAAATAGCGAGAG

GCTTTTG

CAAAAGA

AGTTTTG

CCAGAGG

GGGTA

CAAATC

ATGAAGA

AAGTCTA

CGCTCAC

TGTTATC

TGAAAT

CTGCAATGTGCGAGAAATGACTGAGA

ATTCGTAATCATGGTCATAGC

ATCCCCGGGTACCGAGCTCTACCGTGCAAAATTATTATCA

GGTGGAAACGATACTTGCCCTCTCTATGATAAATGCACTTCCGAGTC

ACAAAACATATAGATGATTA

CGGGTTTTGTTTTATGGAGGT

ATATTACATAACAATCCTC

GCACTCG

CGGGGAT

TTATTTT

ATCTGAA

AACAACATGTTCAGCTA

TTTTGTA

CGGGAGG

CGACTTG

AACCTCC

AACGCGCCTGTTTATCAACAATAGAGGTATTCTAAGA

ACGCGAGG

AGTCCTGAACAAGAAAAA

GATATAGAAGGCTTATCCTA

CATCCTAATTTACGAGCATGTAGAGG

AATCATT

ACCGCGC

CCAATA

TTCATCGTAAACCAATCAAT

TTTTATT

AACAAGCAAGCCGT

CTGTCTTTCCTT

ATCATTC

CAAGAAC

GGGTATT

AAACCAA

GTACCG

Fig.

S9.

Pillar

de

sig

n s

che

ma

tic.

12

Start End Sequence Modified in structure Modification 6[279] 10[266] CGCGACCTGCTCCAACCGCCACCATCAATTCACCCATGCAGG unmodified 2[146] 6[133] ATAAACACCGGAATAAGCATCGTCTCTGCAAGAGAATCTTGA unmodified 6[363] 10[350] TCTTTGACCCCCAGACACTGATATTTAATTACAGCGGCAGTTAATCGAA unmodified 5[35] 6[28] TTAAATCCACGAAGGTGTTAACTTATGCTGGGCTT unmodified 10[370] 2[357] GGAAGAGTTTCTGCGCAACACTAACATCTTAATTTTAGTGAA unmodified 2[125] 6[112] ACTAGAAAAAGCCTCAGCAGCGTAAGCGTATTAAGTCATTAC unmodified 9[567] 5[580] ACATTATCACCATCAAAATCTCCGTCACCGCAAAGACACCAC unmodified 6[174] 10[161] CAGGCGCATAGGCTTTGCTCATTGACCGGGCGAAAGTCGTCT unmodified 9[126] 5[139] CGCGTTTGGGCGCAGACTCCTAATTTACGCCAGTTACAAAAT unmodified 5[518] 1[531] TAGCAAATAGCACCTACCATAATCAGAGTTTTTGGGGTCGAG unmodified 2[104] 6[91] ATCATATGCGTTATCAACAGTTGATGATTATTATTCAAAGCT unmodified 1[280] 9[293] AGAGTTGATTGCAATATTAACAACCAATGTTTTAAATATGCA unmodified 1[238] 9[251] AGACGGGCGCTCAACGGCCAGCCTGTAGGCTTAGAGCTTAAT unmodified 10[244] 2[231] TGACGCTGGCATTCAAAACGATCGTCTGATATCTTTTTTCAA unmodified 10[517] 2[504] GCGTGAGTATTACGTTATCAGTAAACAGGAAGGGTAATTACC unmodified 2[314] 6[301] CGGCTTAGGTTGGGTATTAGAGCCGCCACACCCTCTCATCGC unmodified 1[364] 9[377] TTATAAATTAGTAAAGCAATAATTGTATTAGTTTGACCATTA unmodified 1[343] 9[356] TTCCGAATGAGTAGAGAAGAAATTGTAAAATTCTGCGAACGA unmodified 9[294] 5[307] ACTAAAGTCAGCTCGAGCCACCCCTCAGGTAATTGAGCGCTA unmodified 9[357] 5[370] GTAGATTAAGCAAAGTTTCGTTTTTCATCAAGAAACAATGAA unmodified 5[329] 1[342] ACCCACAAGAATTGAGCCACCTCGTATTCAAACTACTGTTTGATGGTGG Fig. 4A ATACATCTA 9[189] 5[202] GAGAGTACGGATTCTCGAGAGTGGCCTTTAACGTCAAAAATG unmodified 6[384] 10[371] AAGAATACACTAAAACAAACTCAGGAAGAAAATGCGTGACCT unmodified 27[35] 7[48] ACCGCGCTTTTATTGGGTATTAGTAAATGATGCAGACAACAG unmodified 5[476] 1[489] AAAAGAAAACCATCATTGTTTAGACAGGCAGGGCGATGGCCC unmodified 10[328] 2[315] AACTATCGACATCACAGACATTGCTGGTAACAATTTAACCTC Dimers A15 5[455] 1[468] CGCAATAAATCAGTCAATTCAATCCTGAAAAGGGCGAAAAAC unmodified 2[41] 19[55] AGAATCGTTATTTTCGCTCACATTCGTAATCATGGTCATAGC unmodified 10[307] 2[294] TCGCTATTACGGGGTCAGGGATATTACCTTAGAAGTTATATA unmodified 7[49] 25[49] TGCCCGTACACCAGAACGAGTAAACCAACGACTTGACGCGAGG unmodified 1[490] 9[503] ACTACGTATTAAAGGGAGGATAAGGCTACATACAGGCAAGGC unmodified 5[161] 1[174] CAATCCACCTCATTCAAACCCGCCAACACATTAATGAATCGG unmodified 5[287] 1[300] TCAGAGGAACCGCCTGAGGATGCCAGCCCAGCAAGCGGTCCA unmodified 11[56] 17[53] CACCGCTCCGTTTAGCTGAAAGGGTA unmodified 9[147] 5[160] AAGCGAAATAGGTCGGATTAGGAATGGACATATTATTTATCC unmodified 1[301] 9[314] CGCTGGTAGAACAATTAATGATGTTAAATACGGTGTCTGGAA Monomers, Dimers A15 2[83] 6[70] CTTACCAGTATAAAGGTCAGTGTAATAACCCCCTGTAAGGCT unmodified 9[462] 5[475] ATAGTAGTGGAGCAGAATTTTGCACCGTATAACGGAATACCC unmodified 5[56] 1[69] TTTTGCACGGGGTCAACAGAGAACATCGAGCATAAAGTGTAA unmodified

13

9[441] 5[454] AAGGTGGTGAACGGTCTTTCCGAATCAAGGAAACCGAGGAAA unmodified 5[413] 1[426] AGCAGATGTAGCGCAGGAGCGAAGAGTCTGGAACAAGAGTCC unmodified 2[167] 6[154] TAAATAAGGCGTTACAAATATAAAGCCACGGGGTTGGCTGAC unmodified 1[217] 9[230] TGGTTTTTTATTTAAGTCACGCATTAAAAAGAGGTCATTTTT unmodified 10[580] 2[567] TGCGCTAACAGTAGAGAGCAGGTACTATAGATTTTAAAAGAA unmodified 9[231] 5[244] GCGGATGCCAGCTTCCGTACTCCGCCGCCATAAAAACAGGGA unmodified 18[27] 22[8] ATGAAGAGCACTCGACAAAACATATAGATGATTA unmodified 18[54] 11[55] TGAAATAGCCGGACCATTAACGTCCGG unmodified 10[433] 2[420] CCAGTTTCTTGTTGAGTGTGGCGAGTAAGAATTATATTTTCC unmodified 10[76] 2[63] AAACATTGCTGATATCTGGTGGCCCTAAGTGAGGCGCCAACG unmodified 2[503] 6[490] TTTTTTAATGGAAAACTTCTGAACGTCAAACTTTCAACGGCT unmodified 2[335] 6[322] TTTTCGACAACACCGGAACAAGCCCAAAGTAC Fig. 4A ATACATCTA 9[483] 5[496] AATAAATTCAGGTCGCTAAACCCAATGACTGGCATGATTAAG unmodified 6[153] 10[140] CTTCATCAAGAGTAAGGATTAACGTTGGCTCTTCGGAGTCTG unmodified 9[609] 5[622] TCAACGCGTAAAGAATCGGTTGGTAAATAGAAAATTCATATG unmodified 20[51] 20[12] ATCCCCGGGTACCGAGCTCTACCGTGCAAAATTATTATCA unmodified 6[510] 10[497] GGAACGAGGGTAGCAACAGTTATCTACACTGGAACAAGGTGT unmodified 2[608] 6[595] CGCAGAGGCGAATTATACAGTGAAATTAGAGCCTTCGCATAA unmodified 12[48] 16[28] ACCATAAATCAACGAGTTTTGGATAGCGTCAACCC unmodified 9[210] 5[223] TTTTGATTGTGAGCTAGCCCGGGTTGAGCAGCCTTTACAGAG unmodified 10[475] 2[462] TATTGCTAAACTGGAGGGTTGCGCCAGATCAATATTGAGTGA unmodified 14[55] 15[55] ATCCCCCTCAAATGCTTTAGAATACTGCGGAATCGTCATAAA unmodified 10[412] 2[399] TCCTGGGAAGACTCTATCTGCCACGCAAAAGAAACAAACATA unmodified 1[385] 9[398] CGAGATAGTTGTAGGAACCACAAAAGCCTTCGCAAATGGTCA unmodified 5[581] 1[594] GGAATAAAAGGTGAAACGTCACCGCTACGCTTGACGGGGAAA unmodified 2[482] 6[469] TAAATCAATATATGAATCCTGGATAGCACTGTATGGACTAAA unmodified 9[168] 5[181] AACTCCACGGCGGAGTACCAGAATAAATAATAAGAAACGATT unmodified 3[28] 2[42] AGTATGTAGAACCACCAGCAGTTAATTG unmodified 2[62] 4[49] CTCAACAGTAGGGCAAGATAAAGTGCCT unmodified 1[70] 9[83] AGCCTGGACTGATACCGGAAACAGGAAGAAGCAAAGCGGATT unmodified 1[595] 9[608] GCCGGCGCCGCGCTTTTGACGGGGTGAGGAGAAGCCTTTATT unmodified 9[273] 5[286] TCAACATAGGAACGCCCTCAGGAGCCACCACCCTGAACAAAG unmodified 6[468] 10[455] GACTTTTTCATGAGAGTAAATAACAAGAGTCGTTCAAAGCAA unmodified 1[511] 9[524] ATCAAGTCGGGAGCTAAACAGGCTATTTTAGCAAAATTAAGC unmodified 8[27] 4[13] CTGGCTCGAATTACTATAAAGTACCGACA unmodified 5[497] 1[510] ACTCCTTGGCCGGAAATAATGGAGGCCGGAACCATCACCCAA unmodified 6[195] 10[182] ACAGATGAACGGTGAGTGCCGTCCGTGGAAGGCGATGTGAAA unmodified 1[322] 9[335] GAAAATCTCGGCCTCATTGATTTAAAATTCCATATAACAGTT Monomers, Dimers A15 10[223] 2[210] AGGAGAGTGAGATCTTTTCCCCATTGGCAATTGAGATTTCAT unmodified 9[378] 5[391] GATACATCCAAAAAACAACGCCCCTTATTAGCTATCTTACCG unmodified 2[188] 6[175] TTTGAAATACCGACATATCTGACAAACAGCGGATATACAGAC unmodified

14

6[69] 9[62] TGCCCTGACGAGAAATAAACATCCAGCCAGCTTGTTACCCTGACTATTA unmodified 9[504] 5[517] AAAGAATTTGAGAGTCAGCGGTTAGCAAATTACGCAGTATGT unmodified 9[63] 5[76] TAGTCAGATCGCACGTTAATGGTTTTAACCCAGCTACAATTT unmodified 10[454] 2[441] CGAAGTCCGTGAAGAACGAAGTTTTATATTATCAGGTAAATC unmodified 2[356] 6[343] TTTATCAAAATCATTTGCCCGATCACCGTACCGTACGATTAT unmodified 5[371] 1[384] ATAGCAATAGCGTTTTGAGTATCTTTGATCAAAAGAATAGCC unmodified 10[34] 13[48] AGGAGTTCAAAAGAGAACAACACTAATGCAAACAGTTCAGAA unmodified 2[230] 6[217] ATATATTTTAGTTAGAAGGTTGACAGGAGAATAGGGAACCGA unmodified 6[342] 10[329] ACCAAGCGCGAAACAATAGGATATTTTGTCAGCATCGTTAGT Monomers, Dimers A15 2[587] 6[574] TCAATTACCTGAGCCAGGTTTATTATCACCAAAAACGCTGAG unmodified 10[496] 2[483] TATCTCGGATGGGAGTTGCCAGGATTTTGGATTATCAGTACA unmodified 1[553] 9[566] GAACCCTTTGACGACTGCGTGTATTCAACGGTTGTACCAAAA unmodified 5[350] 1[363] CCCAATAATAAGAGAATCAAAAACGTTAACTTGCCATCGGCAAAATCCC unmodified 1[406] 9[419] TCCAGTTTGTCCATCACTCATTGTACCCGTTTAGCTATATTT unmodified 2[566] 6[553] GATGATGAAACAAATAAAGAAAGCCATTATTTTTTAAAGGCC unmodified 6[216] 10[203] ACTGACCAACTTTGATAAGTAGAGTAACGCCAGGGGGTTTTG unmodified 10[559] 2[546] GCGTGTTGAGCTTGTCAGGCAGCACGTAACAGAAACATCAAG unmodified 1[133] 9[146] CCAGTCGGAAAGCGTGCGGGCTGTAGATTAATTCGAGCTTCA unmodified 10[622] 2[609] CCAGCAATACATCATTTCCATAACCACCCCTTTTAAAAATCG unmodified 16[53] 0[42] ATGTTTAGACTGCCAGAGGCGACATAGCAGATCAAATACC unmodified 2[524] 6[511] AACAATTTCATTTGTAGAACCATTACCAAGTGAGACAGCATC unmodified 9[588] 5[601] TTTGCGGAAAGGCCCCAAAAGTTCATTAGTTTATTTTGTCAC unmodified 2[209] 6[196] CTTCTGACCTAAATCAACAGTAGACGATGGTTGATAAAGAGG unmodified 5[602] 1[615] AATCAATATTGACGAACAGTAACACCCGAACGTGGCGAGAAA unmodified 9[35] 10[35] AAAAATCAGGTCTTGTCTTGTCTCCATTATGTTGTCATTGCA unmodified 5[140] 1[153] AAACAGCAAGCGCAACCTTGCCTGACCTGGAAACCTGTCGTG unmodified 5[245] 1[258] AGCGCATAACCACCACTAACAATTTTGACAACAGCTGATTGC unmodified 10[139] 2[126] TAGTGTCAGATGATCGATCGGTAAGAATAAATCTACATAATT unmodified 2[251] 6[238] AACGCGAGAAAACTTAGGAGCACCAGAGCAGGAGGCGCAGAC unmodified 1[259] 9[272] CCTTCACATGGAAAATTCGCAAAATAATTATAATGCTGTAGC unmodified 1[91] 9[104] GAGCTAACTATTAGATTCGCCGGACGACAAAGATTAAGAGGA unmodified 5[119] 1[132] CTAATTTCGTTCCAAAATGAAACGTGGCTCACTGCCCGCTTT unmodified 6[321] 10[308] AACGGAGATTTGTAATTTTCAAAATTTTAAGATGGTTACGCA Fig. 4A ATACATCTA 1[532] 9[545] GTGCCGTCTTTCCTAGCGAAAAAATTAACCTCAGAGCATAAA unmodified 28[55] 27[47] AACAAGCAAGCCGTCCAATA unmodified 1[448] 9[461] CAACGTCGAAGTGTACGCCTGGAATCGACATCAATTCTACTA unmodified 22[48] 2[28] CGGGTTTTGTTTTATGGAGGTCGGGGATCCATATT unmodified 6[237] 10[224] GGTCAATCATAAGGTGTATCATCATCAAACGTTGTGCATCAA unmodified 24[34] 28[15] TTTTGTACAAGAACTTCATCGTAAACCAATCAAT unmodified 6[90] 10[77] GCTCATTCAGTGAACCTATTTTCGGCCTCCAGGCATTTACAT unmodified 6[489] 10[476] ACAGAGGCTTTGAGGGATTTTATTGCCTTGTGCAAGTAAGCG unmodified

15

5[203] 1[216] AAAATAGGCAGGTCTGAAAGGAGATTCAATTGGGCGCCAGGG unmodified 10[160] 2[147] TCACAGCGATGCCACTATTACAACCCTTTGAACCTAATAAGA unmodified 5[434] 1[447] ACCAGAAGTTTGCCTTCCTGAATCAGTGAAGAACGTGGACTC unmodified 9[399] 5[412] ATAACCTCGGTTGAAGACAGCTCGGCATTTTTAAGAAAAGTA unmodified 5[266] 1[279] AACTGAACACCCTCGCCGTCAAACGCTCCGCCTGGCCCTGAG unmodified 5[560] 1[573] AAAGAAACGACTTGATTGCGTGGTTGCTAAAGGGAGCCCCCG unmodified 5[182] 1[195] TTTTGTTGATATTCGTCAGTTAGTAATAGCGGGGAGAGGCGG unmodified 1[427] 9[440] ACTATTAAGGCCACCGATCCGAAAACTAGGGCGCGAGCTGAA unmodified 6[594] 10[581] CCGATATATTCGGTAAAGGCTGGAGACAATGACGAAGGATAT unmodified 1[469] 9[482] CGTCTATAACGGTATCGGACTGAGAGTCTAGCATTAACATCC unmodified 5[539] 1[552] ACATAAATAGAGCCACGTAAATAACGTGAAAGCACTAAATCG unmodified 1[574] 9[587] ATTTAGAAGGGCGCGCAATGCGTCAAATGACCCTGTAATACT unmodified 6[447] 10[434] CCATTAAACGGGTATTTGTCGTAATCGTATAGATGACGGAAA unmodified 10[391] 2[378] CAAGCACTGCACTGGCCGCCTCAATACTACATTATTAAGACG unmodified 10[202] 2[189] TAAAAGATAACGCTTTAAGTTCACGACCGGCAAATTTAATGG unmodified 2[272] 6[259] AAATCCAATCGCAAGATTAGAAGAGCCGCCTCAGATGTTACT unmodified 6[426] 10[413] TAATGCCACTACGAGTTAGCGAATCATATGTTGTGTTCGCCA unmodified 9[546] 5[559] GCTAAATCCGTTCTAATAATATGGGAATGGTGGCAACATATA unmodified 2[545] 6[532] AAAACAAAATTAATTATTTGCAGCAAAACTAAAGGGTCACCC unmodified 1[196] 9[209] TTTGCGTCCAGTCAGGGTAACAACCCGTCCTTTAATTGCTCC unmodified 6[405] 10[392] AACCTAAAACGAAAATTCCACTAATCAGCAGGCTACTGTTAT unmodified 2[398] 6[385] GCGATAGCTTAGATCATTTTGCATAGCCCTGTAGCGAGGCAA unmodified 10[181] 2[168] ATGCTGAATTTCGCGGGGGATCATTCTGTCAATCACGTGTGA unmodified 1[154] 9[167] CCAGCTGGAGATAGGCCAGCTTAATGGGCCAGACCGGAAGCA unmodified 9[315] 5[328] GTTTCATTCGCATTGGGATAGCCGCCTCAGAGAT Dimer, Fig. 4 A, E A15 9[315] 5[328] GTTTCATTCGCATTGGGATAGCCGCCTCAGAGAT Dimer, Fig 1A A15, ATTO647N 10[538] 2[525] AAAATCAAAACTCACGAAGTGCGTTAGATCAAAATTACATTT unmodified 1[112] 9[125] GTTGCGCACAGACAACTGTTGCGTGCATAAGACTTCAAATAT unmodified 10[349] 2[336] CAAGACCCAGAAATAAGAACTAAATCCTAGGTCTGAGAGACTACC Dimer A15 2[440] 6[427] GTCGCTATTAATTACATCATATTTAGCGCTAAAGTAAATACG

unmodified

9[336] 5[349] TATAACAGTTGATTCCCACGTTAAACCCATGGAACCAGAGTTAAG Fig. 4A ATACATCTA 6[111] 10[98] CCAAATCAACGTAACTGAAACTTTGAGGATTCAGGTGAGTAT unmodified 10[286] 2[273] TCAATGGGTTCAGGTCATGCGCAGGAAAATAGATACTGATGC unmodified 9[420] 5[433] TCATTTGGCATGTCTAACGATTCAGACTAGCCGAACAAAGTT unmodified 9[525] 5[538] AATAAAGTGCCGGAGGAACAATCACCAGCGTAGAAAATACAT unmodified 6[258] 10[245] TAGCCGGAACGAGGTTTAGTATGGCCTTTGCCAAGTGATGTC unmodified 9[105] 5[118] AGCCCGACTGCCAGATGAAAGTCATACACGTCTTTCCAGAGC unmodified 1[175] 9[188] CCAACGCAAAGGGAGTGCTGCGAACAAAACAGGTCAGGATTA unmodified 16[27] 0[14] TCGTTTAGCTTTTGTATAAATATTGGGCTAGATATAGAGTCG unmodified 10[601] 2[588] CGACCAGGAGAACGCTGGGGATAATGCGGATGAATATTCATT unmodified 6[573] 10[560] GCTTGCAGGGAGTTCACGTTGAATATGAAAGCGGCGGAAACT unmodified

16

2[293] 6[280] ACTATATGTAAATGATACATTACCCTCAAACCGCCCGAAATC unmodified 2[377] 6[364] CTGAGAAGAGTCAATAAAAGTTGCCATCCACCAGTACACTCA unmodified 10[118] 2[105] CTCAAACATCGGGTCTGCGCAATATTTTTGAGAGCGTTTAGT unmodified 26[47] 27[34] GATATAGAAGGCTTATCCTAAATCATT unmodified 4[48] 29[54] TGAGTATAAGATTAGTTGCTAAACCTCCGTACCG unmodified 5[98] 1[111] TAACGAGTGGCTTTGCCACGCTGAATGGCTCACATTAATTGC unmodified 10[265] 2[252] TGAGTATCTGCATACTTTATTTACCTACACTAATAGACAAAG unmodified 2[419] 6[406] CTTAGAATCCTTGACACCAGAGTTTTCACCTCATAAGGCACC unmodified 6[300] 10[287] CTGATAAATTGTGTACCCTCAATTTTTTCAACAGTTTGGAGG unmodified 0[41] 23[48] GAACGATCCACACAACATACGTGTTATCATCTGAA unmodified 10[97] 2[84] TATCTTACTGTTTCAAGCGCCTCTTTAACCGCCTGACAAATT unmodified 9[84] 5[97] GCATCAAGACAGTACGGAACCACAGGAGAATCTTACCAACGC unmodified 5[308] 1[321] ACAAATATCCTTGCCCCAGCAGGCGAA Fig. 4A ATACATCTA

5[308] 1[321] ACAAATATCCTTGCCCCAGCAGGCGAA Fig. 4E TTTCCCACCGCTCG GCTCAACTGGG

5[77] 1[90] TATCCTGTGTACTGATTAACATGCGCGAGGTGCCTAATGAGT unmodified 5[224] 1[237] AGAATAACAGCATTATCTAAAAAATGGATCTTTTCACCAGTG unmodified 5[392] 1[405] AAGCCCTTTTCGGTCGGAACAATTAACCGGGTTGAGTGTTGT unmodified 9[252] 5[265] TGCTGAATCGCGTCCCGCCACCCACCAGTAGACGGGAGAATT unmodified 6[531] 10[518] TCAGCAGCGAAAGAATAGAAAGAGGGTAAGAGGTTACGAGCA unmodified 6[132] 10[119] CAAGAACCGGATATAGGCTGATCGTAACGGAAGGGGACCGTA unmodified 6[552] 10[539] GCTTTTGCGGGATCAATTGCGAGCTGATTTAACTCAAACAGC unmodified 2[461] 6[448] ATAACCTTGCTTCTATGATGGAGCGACAAGACGTTGAAGTTT unmodified 9[2] 12[8] TGGGAAGAAAAATCTACGTCAGGTAGAAAGAT 5'-Biotin 25[5] 24[35] AACGCGCCTGTTTATCAACAATAGAGGTATTCTAAGACGGGAGG 5'-Biotin 17[5] 16[11] ACCAAAATAGCGAGAGCCAGACGACG 5'-Biotin 11[5] 10[11] GAGTCTGAGCAAAAAGGAGTATCAAT 5'-Biotin 7[9] 8[8] TTTAATCATTGTATTATACCAGTCA 5'-Biotin 5[5] 5[34] AGTAATTCTGTCCAGACGACGACAAAAGCC 5'-Biotin 3[2] 2[9] AGAGGCATTTTCGAGCCAGTAATAAGTAACAACGCCAACATGTAA 5'-Biotin 1[9] 8[28] CAAATATTCCATGAAGGTTTATAATTGAAGGTAGTTATCATTAAGAA 5'-Biotin 13[2] 9[34] GTTGAGATTTAGGAATACCACATTCAATTATTATAATAAAACGAACT 5'-Biotin 23[2] 18[15] ATATTACATAACAATCCTCCAAATC 5'-Biotin 29[9] 6[14] CTGTCTTTCCTTTTTAATT 5'-Biotin 19[9] 18[28] CTGCAATGTGCGAGAAATGACTGAGAAAGTCTA 5'-Biotin 15[9] 14[15] GCATAGTAAGAGCAACACTATCATCATACATAACGCCAAAAGGAAT 5'-Biotin 21[5] 21[51] GGTGGAAACGATACTTGCCCTCTCTATGATAAATGCACTTCCGAGTC 5'-Biotin 27[2] 26[10] CATCCTAATTTACGAGCATGTAGAGGAGTCCTGAACAAGAAAAA 5'-Biotin 6[27] 24[11] GAGATGGATCATTCAACAACATGTTCAGCTA unmodified 6[615] 10[602] ACAACCATCGCCCATAATTGTTTCAAAACAGACCTAACGCCG unmodified

17

Table S1. List of unmodified and modified oligonucleotides from the 5’ to the 3’ used for the different DNA pillar structures sketched in Fig. 1A, Fig. 4A and E. The nomenclature follows the numbering used in Fig. S9.

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