DNA nanotechnology as a tool to manipulate lipid bilayer membranes
Meng, Zhuojun
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Chapter 4
DNA Replacement and Hybridization Chain
Reaction on the Surface of Liposome Membrane
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4.1 Introduction
Synthetic biology and cell surface engineering techniques vitro and in-vivo have resulted in novel tools for the development of membrane biology,1 offering promising membrane-based devices that may enable new
types of artificial tissues,2,3 biosensors,4,5 drug delivery approaches,6,7 3D
bio-printing and the study of lipid metabolism.8 To boost the development
of these technologies, there is a growing need to enhance surface engineering techniques of membranes under in-vitro and in-vivo conditions with particular emphasis on exploiting artificial surface receptors9 and
75 designing novel biomaterials guided by natural processes,10 such as
self-assembling peptides, proteins and DNA oligonucleotides. Especially the latter class of biomacromolecules is very appealing to fabricate complex architectures because the sequence specific base pairing of oligonucleotides allows the prediction of the resulting structure based on the sequence composition, qualifying nucleic acids as indispensible building blocks in soft matter nanotechnology.11 In conjunction with
advances in solid phase DNA synthesis methods,12 a plethora of
programmed 2- and 3-dimensional self-assembled architectures was achieved.13,14
The facile chemical modification of oligonucleotides with hydrophobic anchors also allowed the fabrication of DNA-based functional membranes.15 In the context of liposomes, DNA hybridization-induced
vesicle aggregation,16 and fusion were realized.17 Photoresponsive
DNA-lipid assemblies, fabricated by either anchoring DNA with a azobenzene moiety18 or hybridization of a photosensitizer mediated the cargo release
from liposomes.19 While these functions relied on simple DNA amphiphiles
that were inserted in the membrane, vesicle deformation and even destruction of these containers was achieved with immobilizing and polymerizing more complex DNA origami structures.13,20 Further extension
of these concepts led to a DNA-based atomistically determined molecular valve capable of controlling transport of small molecules across a biological membrane.21
In this chapter, we implement such membrane engineering related DNA nanotechnology on the surface of a phospholipid bilayer. We performed studies to establish the anchoring of DNA amphiphiles in such a bilayer, subsequent hybridization, strand replacement and DNA hybridization chain reaction (HCR). For that purpose, vesicles served as a model system. Previously, hybridization of DNA on a liposome surface was demonstrated,22 however, strand replacement and DNA HCR, to the best of
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4.2 Results and Discussion
4.2.1 Design and definition of lipid-DNA
For surface anchoring of oligonucleotides, we employed lipid-modified DNA,23 consisting of a hydrophobic alkyl chain and an ethyne function
attached to the nucleobase, i.e. at the 5-position of uracil (Scheme 4.1A). The incorporation of the hydrophobic building blocks was achieved as phosphoramidites by solid phase synthesis employing an automated DNA synthesizer and a previously established procedure.24 Due to this
convenient incorporation method, multiple hydrophobic nucleotides can be introduced into the same oligonucleotide at any desired position allowing to tune the interaction with phospholipid membranes. Here, we chose four lipid-modified deoxyuridine units attached either to the 3’- or to 5’-end of the oligonucleotide sequences, which are comprised of 18 or 28 nucleotides (Scheme 4.1B-D).
Scheme 4.1 Schematic representation of structures: (A) Chemical structure of lipid-modified
77 They are abbreviated as UxTy, where x represents the number of lipid-modified uracils at the terminus while y denotes the overall number of nucleotides of the sequence (Table 4.1). The four consecutive hydrophobic anchoring units guarantee stable incorporation into a phospholipid membrane of vesicles for at least 24 h as proven by a fluorometric assay, which was described in Chapter 2.24
Table 4.1 Sequences and modifications of DNA.
DNA Sequence ( 5’→3’) * U4T-18 5’-UUUUGCGGATTCGTCTGC-3’ CrU4T-18 5’-GCAGACGAATCCGCUUUU-3’ U4T-28 5’-UUUUAATTGGGTGCGGCTTAGGATCTGA-3’ C488 5’-GCAGACAGGTCCGC-3’-ATTO488 C594 5’-GCAGACAGGTCCGCGTTTGT-3’-ATTO594 20-mer 5’-ACAAACGCGGATTCGTCTGC-3’ M1 5’-GTGCGGCTTAGGATCTGATGAAA CTCAGATCCTAAGCCGCACCCAATT-3’ M1-FAM 6-FAM-5’-GTGCGGCTTAGGATCTGATGAAACTCA GATCCTAAGCCGCACCCAATT-3’ M2 5’-GTTTCATCAGATCCTAAGCCGCACAAT TGGGTGCGGCTTAGGATCTGA-3’ M2-Cy3 Cy3-5’-GTTTCATCAGATCCTAAGCCGCACAAT TGGGTGCGGCTTAGGATCTGA-3’
*: U represents the modified uracil base. ATTO594, ATTO488, 6-FAM, Cy3 represent fluorescent dyes covalently bound to the DNA oligonucleotides.
4.2.2 DNA hybridization and replacement on the surface of liposomes
Firstly, U4T-18 was stably anchored into the membrane of vesicles (diameter 120 nm) by in situ modification, i.e. by addition of a U4T-18 solution to plain liposomes (DOPC:DOPE:Cholesterol, 2:1:1, molar ratio). Followed by 1h incubation at 50 °C. In situ modification spontaneously78
occurred due to the hydrophobic units part of lipid-DNA piercing into the interior of the phospholipid bilayer. Then two Fluorescence Resonance Energy Transfer (FRET) systems, Rhodamine Rh-DHPE/C594 and C488/Rh-DHPE, were used to demonstrate the availability of the anchor sequence for hybridization and DNA replacement (Fig 4.2A).
Fig. 4.2 (A) Schematic representation of DNA replacement on the surface of liposomes. (B)
Fluorescence spectra (λEX = 470 nm) of FRET system with U4T-18/Rh-DHPE/C594. FRET is
achieved when C594 hybridizes with U4T-18 to bring the donor Rh closer to the acceptor C594 (dashed purple line). Afterwards, C594 was peeled off from U4T-18 by hybridizing with 20-mer (black line). Disruption of liposomes by addition of Triton X-100 (0.3% (v/v)) results in termination of FRET (dashed red line). (C) After C594 was removed (black line Fig. 4.2B), U4T-18 remained on the liposome and maintained the ability to hybridize with C488, which leads to FRET between donor C488, and acceptor Rh-DHPE (black line). Liposome disassembly after the addition of 0.3% (v/v) Triton X-100 results in an increase of C488 donor emission (dashed red line).
In the Rh-DHPE/C594 system, ATTO594 was covalently attached to the 3’ end of a 20mer DNA (C594) to act as an acceptor. In parallel, rhodamine-functionalized phospholipid (Rh-DHPE) was incorporated into the liposomal bilayer to function as a donor (Fig 4.2A, step 1). As demonstrated by a clear emission peak at 624 nm (C594, acceptor fluorescence peak), hybridization only occurred upon mixing of C594 with
79 U4T-18-grafted Rh-DHPE-containing vesicles, positioning both dyes sufficiently close to each other to achieve FRET (Fig 4.2B, dashed purple line). Afterwards, a 20-mer DNA oligonucleotide that is fully complementary to C594 was introduced to peel off C594 from U4T-18 (Fig 4.2A, step 2), which resulted in a higher signal of the donor emission of Rh-DHPE (592 nm) and lowering of the acceptor emission (C594, 624 nm) signal (Fig 4.2B, black line). Then, a 14-mer DNA which is complementary to U4T-18 was covalently attached to ATTO488 and hybridized with free U4T-18 (Fig 4.2A, step 3). In this case, C488 acted as donor for Rh-DHPE which forms a second FRET system: C488/Rh-DHPE. Disruption of liposomes by addition of Triton X-100 resulted in an increased donor peak (C488, 520 nm) and slightly decreased acceptor peak (Rh-DHPE, 592 nm) (Fig 4.2C), confirming that replacement by hybridization on the surface of the vesicles was successfully achieved.
Fig. 4.3 Absence of FRET when liposomes are decorated with CrU4T-18. (A) Fluorescence
spectra (λEX = 470 nm) of non-FRET system with CrU4T-18/Rh-DHPE/C594 (dashed purple
line). After 1h incubation, 20-mer was added to the system (black line). Disruption of liposomes by addition of Triton X-100 (0.3% (v/v)) results in termination of FRET (dashed red line). (B) Fluorescence spectra of non-FRET system with CrU4T-18/C488/Rh-DHPE before (black line) and after (dashed red line) addition of Triton X-100.
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In the control experiments involving CrU4T-18-liposomes, which cannot hybridize with C594 or C488, similar emission spectra were obtained before and after liposomal disruption, indicating that no FRET occurred in the absence of complementary anchoring units on the vesicle surface (Fig. 4.3).
Moreover, native polyacrylamide gel electrophoresis (PAGE) was performed to demonstrate DNA replacement in buffer in the absence of liposomes (Fig. 4.4). The results showed that both C488 and C594 hybridize with U4T-18 (lane 4 and lane 5, respectively) and that the 20-mer can efficiently peel off C594 from U4T-18 (lane 6). After C594 was removed from U4T-18 by 20-mer, C488 hybridized with free U4T-18 (lane 7).
Fig. 4.4 PAGE of DNA replacement in buffer (M is maker). The sample run from left to right,
Lane 1: C488; Lane 2: C594; Lane 3: 20-mer; Lane 4: U4T-18 + C488; Lane 5: U4T-18 + C594; Lane 6: C594 was peeled off from U4T-18 by 20-mer; Lane 7: C488 hybridized with free U4T-18 from Lane 6; Lane 8: C488 + 20-mer; Lane 9: C594 + 20-mer. The excitation with UV was at 366 nm.
4.2.2 DNA hybridization chain reaction (HCR) on the surface of
liposomes
Since initiation of HCR from a lipid membrane was not demonstrated before we first established a HCR protocol for decorating the rim of liposomes with a DNA layer. Membrane anchor U4T-28, a 28-mer
lipid-81 DNA with 4 modified lipid bases, was introduced to liposomes. Next, hairpin strands M1 (partially complementary to U4T-28) and M2 (partially complementary to M1) were added. Hybridization of M1 to U4T-28 results in liberation of its loop that subsequently can hybridize with M2.25 Opening
of the M2 hairpin exposes a sequence that binds to a new M1 monomer from the solution. In turn, opening of the M1 hairpin exposes a sequence that can bind new M2. This effectively triggers the “supramolecular polymerization” of M1 and M2 with surface anchor U4T-28 as initiator (Fig. 4.5).
Fig. 4.5 Schematic representation of lipid-DNA initiated HCR . a’, b’, and c’ are regions which
are complementary to regions a, b, and c, respectively. Hairpin M1 can be unfolded by hybridization with initiator U4T-28 or open M2, while hairpin M2 can be unfolded by hybridization with open M1, resulting in growing DNA strands.
Agarose gel electrophoresis analysis was used to prove U4T-28 initiated HCR with M1 and M2 (Fig. 4.6). The hairpin sequences M1 and M2 (Fig. 4.6, lane 1 and 2, respectively) did not hybridize in the absence of U4T-28 (Fig.
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4.6, lane 3). The chain length of the resulting duplex DNA is inversely related to the initiator concentration (Fig. 4.6, lane 6-8).
Fig. 4.6 Agarose gel electrophoresis analysis of DNA HCR. Lane 1: M1; lane 2: M2; lane 3:
M1+M2; lane 4: M1+U4T-28; lane 5: M2+U4T-28; lanes 6–8: three different molar ratios of initiator (1:1:1, 1:1:0.5, 1:1:2, M1:M2:U4T-28).
To prove DNA extension on the surface of liposomes, M2 was labeled with a fluorophore (Cy3). Firstly, U4T-28 was incubated with liposomes for 30min at 50 °C. PTHK polysulfone membrane filters (100 kDa) were used to remove unincorporated U4T-28 by centrifugation. M1 and M2-Cy3 (two equivalents in relation to U4T-28), were added to the system at room temperature for 1 h, after which again a 100 kDa molecular weight cut off filter was used in a centrifugation step to remove free M1 and M2-Cy3. Afterwards, fluorescence intensity of the supernatant containing liposomes was measured. The fluorescence spectra were recorded in the range of 530-620 nm with excitation at 520 nm for M2-Cy3 detection. A clear emission peak at 566 nm (Cy3) only occurs upon chain reaction on the surface of the liposomes (Fig. 4.7B, dashed red line), whereas in the absence of M1 (Fig. 4.7B, dashed blue line) or U4T-28 (Fig. 4.7B, black line), negligible Cy3 signals were observed.
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Fig. 4.7 DNA HCR on the surface of liposomes. (A) Schematic representation of HCR for
decorating the outer layer of liposomes with a DNA shell. Liposomes (DOPC/DOPE/Chol = 2:1:1 mol%) decorated with U4T-28, and incubated with M1 and M2-Cy3 for 1 h, lead to DNA HCR (I). After centrifugation and filtration, the fluorescence of supernatant containing liposomes was measured. (B) Fluorescence spectra of Cy3 in HCR system (I, dashed red line). In the absence of M1 (II, dashed blue line) or U4T-28 (III, black line), no fluorescence was observed.
4.3 Conclusion
In this chapter, we invented new concepts for the DNA functionalization of a liposome surfaces. We explored DNA hybridization and the dynamic exchange of DNA sequences on the surface of liposomes with two FRET systems. As an anchoring unit a DNA amphiphile was utilized. Hydrophobic units were incorporated into nucleobases, which pierce into the interior of the phospholipid bilayer. DNA hybridization on the surface of liposomes was proved by FRET between C594 (acceptor) and U4T-18-grafted Rh-DHPE-containing (donor) vesicles, positioning both dyes sufficiently close to each other to achieve FRET. After that, a 20-mer DNA oligonucleotide that is fully complementary to C594 was introduced to peel off C594 from U4T-18, which was confirmed by the increase of donor signal. Then, C488 hybridized with free U4T-18 acted as a donor for Rh-DHPE. The results demonstrated that the hybridization process can be designed to be
84
reversible allowing exchange of surface functionalities by simple addition of DNA sequences. Finally, a DNA based amplification process was performed atop of the liposome enabling the multiplication of surface functionalities from a single DNA anchoring unit. A DNA probe, M2-Cy3, was employed to detect the DNA HCR on the liposome’s surface. Compared with control experiments, which were lacking DNA oligonucleotide monomer for HCR or of the initiator, a significant stronger fluorescence intensity of Cy3 was observed which can only be rationalized when multiplication of DNA occurs on surface of liposomes. The hybridization chain reaction preformed in this chapter allows accumulation of multiple cargoes or signals on the liposomal surface by using anchored single DNA strands. The experiments shown in this chapter significantly extend the functionality of liposomal system regarding loading and decorating the vesicle surface. This might be exploited in the fields of drug delivery or diagnostics.
4.4 Experimental Section
4.4.1 Materials
Cholesterol (Chol), 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE) and 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) were purchased from Avanti Polar Lipids (Alabaster, USA) (purity >99 %) and used without further purification. Headgroup-labeled phospholipid, lissamine rhodamine B 1,2-dihexadecanoyl-sn-glycero-3-phosphoethanolamine (triethylammoni -um salt) (Rh-DHPE) was purchased from Invitrogen (Amsterdam, Netherlands), and used as received. PTHK polysulfone membrane filters with a NMWL of 100 kDa were purchased from Sigma-Aldrich. The DNA-dye conjugates C488, C594 and M2-Cy3 were purchased from Biomers.net GmbH (Ulm, Germany). Triton X-100 (10% in water) was purchased from Sigma-Aldrich (St. Louis, United States). Anhydrous CHCl3 was acquired
from Acros Organics (Geel, Belgium) and stored over molecular sieves. For all experiments, ultrapure water (specific resistance > 18.4 MΩ cm) was obtained by a Milli-Q water purification system (Sartorius).
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4.4.2 Synthesis and characterization of amphiphilic oligonucleotides
Fig. 4.8 MALDI-TOF mass spectra of (A) U4T-18, (B) U4T-28 and (C) CrU4T-18.
Fig. 4.9 RPC HPLC analysis of purified lipid-DNAs: (A) U4T-18, (B) U4T-28 and (C) CrU4T-18.
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The amphiphilic oligonucleotides were synthesized and purified as reported previously (see Chapter 2).23,26 The identity and purity of the
oligonucleotides were confirmed by RPC-HPLC and MALDI-TOF mass spectrometry (Fig. 4.8 and Fig. 4.9).
4.4.2 Preparation and characterization lipid-DNA liposomes
Firstly, chloroform was removed from lipid mixture (DOPC:DOPE: Cholesterol, 2:1:1, molar ratio) by evaporation under an air stream and then under vacuum overnight. The dried lipid mixture was dissolved in an aqueous PBS buffer (150 mM NaCl, 15 mM K2HPO4, 5 mM KH2PO4) by 5
cycles of vortexing and freeze-thawing. Subsequently, the sample was dispersed by extruding 21 times using an extruder and 100 nm polycarbonate membranes (Whatman) to obtain large unilamellar vesicles (LUVs), after which the liposomes with lipid-DNA were incubated at 50°C for 1h. All lipid-DNA liposomes were used within one day, and had an average diameter of around 120 nm as determined by DLS (ALV/CGS-3 ALV-Laser Vertriebsgesellschaft mbH, Langen, Germany). The molar ratio between lipid and U4T-18 was 500:1.
4.4.3 DNA replacement on liposomes measured by Fluorescence
Resonance Energy Transfer (FRET) assay
U4T-18 was incorporated in Rh-DHPE/(DOPC+DOPE) (3:97 molar ratio) liposomes to obtain U4T-18 liposomes with a lipid to U4T-18 ratio of 500:1. Subsequently, an aliquot of these liposomes was mixed with a small amount of Cr-ATTO488 or CrATTO594 such that [U4T-18] = [Cr-ATTO488] = [Cr-ATTO594] = 0.906 μM and with a final lipid (DOPC+DOPE) concentration of 0.45 mM. Fluorescence emission spectra of donor/acceptor, CrATTO488/Rh-DHPE or Rh-DHPE/CrATTO594 in the 500–700 nm region, were recorded with excitation at 470 nm using a SpectraMax M3 (Molecular Devices) fluorescence spectrophotometer. Measurements were carried out at a constant temperature of 25 °C.
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4.4.4 Native polyacrylamide gel electrophoresis to detect DNA
replacement
Native polyacrylamide gel electrophoresis was performed using a 15% gel made with TBE buffer (90 mM Tris, 89 mM boric acid, 2 mM EDTA, pH 8.0) and run at 120 V for 100 min.
4.4.5 Agarose gel electrophoresis to monitor HCR
The 2 % agarose gel was prepared by using SB buffer (10 mM NaOH, pH adjusted to 8.5 with boric acid) 27 and run at 90 V for 3 h. After
electrophoresis, the agarose gel was stained with ethidium bromide and visualized under UV light.
4.4.6 DNA hybridization chain reaction on the surface of liposomes
After incubation of 0.45 mM liposomes with 0.906 μM U4T-28 at 50 °C for 30 min, PTHK polysulfone membrane filters (100 kDa) were used to remove unincorporated U4T-28 by centrifugation at 1000 rpm (rotor: FA-45-18-11) for 30 min. M1 and M2-Cy3 (two equivalents in relation to U4T-28) were added to the system at RT for 1 h, after which again a 100 kDa molecular weight cut off filter was used in a centrifugation step to remove free DNA (1000 rpm, 30 min). After re-suspension and centrifugation, the supernatant was washed twice with PBS buffer. Afterwards, the fluorescence spectra were recorded in the range of 530-620 nm with excitation at 470 nm for M2-Cy3, using a SpectraMax M3 (Molecular Devices) fluorescence spectrophotometer. Measurements were carried out at 25 °C.Author contributions
Meng Z designed, conducted the experiments and performed data analysis. Meng Z and Yang J prepared the manuscript. Liu Q synthesized lipid-DNA. Kros A and Herrmann A supervised the project.
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References
1. Saeui, C. T.; Mathew, M.P.; Liu, L.; Urias, E.; Yarema, K. J.; Cell Surface and Membrane Engineering: Emerging Technologies and Applications. J. Funct. Biomater. 2015, 6, 454-485. 2. Cukierman, E.; Pankov, R.; Stevens, D. R.; Yamada, K. M.; Taking cell matrix adhesions to the third dimension. Science 2001, 294, 1708-1712.
3. Revzin, A.; Rajagopalan, P.; Tilles, A. W.; Berthiaume, F.; Yarmush, M. L.; Toner, M.; Designing a hepatocellular microenvironment with protein microarraying and poly(ethylene glycol) photolithography. Langmuir 2004, 20, 2999-3005.
4. Shear, J. B.; Fishman, H. A.; Allbritton, N. L.; Garigan, D.; Zare, R. N.; Scheller, R. H.; Single cells as biosensors for chemical separations. Science 1995, 267, 74-77.
5. Rider, T. H.; Petrovick, M. S.; Nargi1, F. E.; Harper, J. D.; Schwoebel, E. D.; Mathews1, R. H.; Blanchard, D. J.; Bortolin, L. T.; Young, A. M.; Chen, J.; Hollis, M. A.; A B cell-based sensor for rapid identification of pathogens. Science 2003, 301, 213-215.
6. Yang, J.; Shimada, Y.; Olsthoorn, R. C. L.; Snaar-Jagalska, B. E.; Spaink, H. P.; Kros, A.; Application of Coiled Coil Peptides in Liposomal Anticancer Drug Delivery Using a Zebrafish Xenograft Model. ACS Nano. 2016, 10, 7428-7435.
7. Yang, J.; Bahreman, A.; Daudey, G.; Bussmann, J.; Olsthoorn, R. C. L.; Kros, A.;.; Drug Delivery via Cell Membrane Fusion Using Lipopeptide Modified Liposomes. ACS Cent. Sci.
2016, 2, 621-630.
8. Han, G. S.; O'Hara, L.; Carman, G. M.; Siniossoglou, S.; An unconventional diacylglycerol kinase that regulates phospholipid synthesis and nuclear membrane growth. J. Biol. Chem.
2008, 283, 20433-20442.
9. Mrksich, M.; What can surface chemistry do for cell biology? Curr. Opin. Chem. Biol. 2002, 6, 794-797.
10. Zhang, S.; Fabrication of novel biomaterials through molecular self-assembly. Nat.
Biotechnol. 2003, 21, 1171-1178.
11. Seeman, N. C.; DNA in a material world. Nature 2003, 421, 427-431.
12. Caruthers, M. H.; Gene synthesis machines: DNA chemistry and its uses. Science 1985, 230, 281-285.
13. Kocabey, S.; Kocabey, S.; Kempter, S.; List, J.; Xing, Y.; Bae, W.; Schiffels, D.; Shih, W. M.; Simmel, F. C.; Liedl, T.; Membrane-Assisted Growth of DNA Origami Nanostructure Arrays.
89
14. Todhunter, M.E.; Jee, N. Y.; Hughes, A. J.; Coyle, M. C.; Cerchiari, A.; Farlow, J.; Garbe, J. C.; LaBarge, M. A.; Desai, T. A.; Gartner, Z. J.; Programmed synthesis of three-dimensional tissues. Nat. Methods 2015, 12, 975-981.
15. Yoshina-Ishii, C.; Boxer, S.G.; Arrays of Mobile Tethered Vesicles on Supported Lipid Bilayers. J. Am. Chem. Soc. 2003,125, 3696-3697.
16. Jakobsen, U.; Simonsen, A.C.; Vogel, S.; DNA-Controlled Assembly of Soft Nanoparticles. J.
Am. Chem. Soc. 2008, 130, 10462-10463.
17. Stengel, G.; Zahn, R.; Höök, F.; DNA-Induced Programmable Fusion of Phospholipid Vesicles. J. Am. Chem. Soc. 2007, 129, 9584-9585.
18. Hernández-Ainsa, S.; Ricci, M.; Hilton, L.; Aviñó, A.; Eritja, R.; Keyser, U. F.; Controlling the Reversible Assembly of Liposomes through a Multistimuli Responsive Anchored DNA. Nano
Lett. 2016, 16, 4462-4466.
19. Rodríguez-Pulido, A.; Kondrachuk, A. I.; Prusty, D. K.; Gao, J.; Loi, M. A.; Herrmann, A.; Light-Triggered Sequence-Specific Cargo Release from DNA Block Copolymer–Lipid Vesicles.
Angew. Chem. Int. Ed. 2013, 52, 1008-1012.
20. Czogalla, A.; Czogalla, A.; Kauert, D. J.; Franquelim, H. G.; Uzunova, V.; Zhang, Y.; Seidel, R.; Schwille, P.; Amphipathic DNA Origami Nanoparticles to Scaffold and Deform Lipid Membrane Vesicles. Angew. Chem. Int. Ed. 2015, 127, 6601-6605.
21. Burns, J. R.; Seifert, A.; Fertig, N.; Howorka, S.; A biomimetic DNA-based channel for the ligand-controlled transport of charged molecular cargo across a biological membrane. Nat.
Nanotechnol. 2016, 11, 152-156.
22. Kurz, A.; Bunge, A.; Windeck, A.; Rost, M.; Flasche, W.; Arbuzova, A.; Strohbach, D.; Müller, S.; Liebscher, J.; Huster, D.; Herrmann, A.; Lipid-Anchored Oligonucleotides for Stable Double-Helix Formation in Distinct Membrane Domains. Angew. Chem. Int. Ed. 2006, 45, 4440-4444.
23. Anaya, M.; Kwak, M.; Musser, A. J.; Mullen, K.; Herrmann, A.; Tunable hydrophobicity in DNA micelles: design, synthesis, and characterization of a new family of DNA amphiphiles.
Chem. Eur. J. 2010, 16, 12852-12859.
24. Meng, Z.; Yang, J.; Liu, Q.; de Vries J. W.; Gruszka, A.; Rodríguez-Pulido, A.; Crielaard, B J.; Kros, A.; Herrmann, A.; Efficient Fusion of Liposomes by Nucleobase Quadruple-Anchored DNA. Chem. Eur. J. 2017, DOI: 10.1002/chem.201701379.
25. Dirks, R. M.; Pierce, N. A.; Triggered amplification by hybridization chain reaction. Proc.
Natl. Acad. Sci. U.S.A. 2004, 101, 15275-15278.
26. Kwak, M.; Minten, I. J.; Anaya, D. M.; Musser, A. J.; Brasch, M.; Nolte, R. J. M.; Müllen, K.; Cornelissen, J. J. L. M.; Herrmann, A.;Virus-like Particles Templated by DNA Micelles: A General Method for Loading Virus Nanocarriers. J. Am. Chem. Soc. 2010, 132, 7834-7835. 27. Brody, J. R.; Kern, S. E.; Sodium boric acid: a Tris-free, cooler conductive medium for DNA electrophoresis. BioTechniques 2004, 36, 214-216.
