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DNA Nanotechnology allows the synthesis of nanometer sized objects that can be site specifically functionalized with a large variety of materials. However, many DNA structures need a higher ionic strength than that in common cell culture buffers or in bodily fluids to maintain their integrity and can be degraded quickly by nucleases. The aim of this dissertation was to overcome this deficiency with the help of cationic PEG-poly-lysine block copolymers that can electrostatically cover the DNA nanostructures to form “DNA origami polyplex micelles” (DOPMs). This straightforward, cost-effective and robust route to protect DNA-based structures could therefore enable applications in biology and nanomedicine, where un-protected DNA origami would be degraded. Moreover, owing to high polarity, the DNA-based structures are restricted to the aque-ous solution based buffers only. Any attempt to change the favorable conditions, leads to the distortion of the structures. In this work it was demonstrated that, by using the polyplex micellization strategy, the organic solubility of DNA origami structures can be improved. The strategy was also extended to functional ligands that are otherwise not soluble in organic solvents. With this strategy, it is now also possible to perform organic solution reactions on the DNA-based structures, opening up the possibility to use hydro-phobic organic reagents to synthesize novel materials. The polyplex micellization strategy therefore presents a cheap, robust, modular, reversible and versatile method to not only solubilize DNA structures in organic solvents but also improve their stability in biological environments. A third project was based on the possibility to synthesize complementary sequences to single-stranded gap regions in the DNA origami scaffold cost-effectively by a DNA polymerase rather than by a DNA synthesizer. For this purpose, four different wireframe DNA origami structures were designed to have single-stranded gap regions. The introduction of flexible gap regions resulted in fully collapsed or partially bent structures due to entropic spring effects. These structures were also used to demonstrate structural transformations with the help of DNA polymerases, expanding the collapsed bent structures to straightened tubes. This approach presents a powerful tool to build DNA wireframe structures more material-efficiently, and to quickly prototype and test new wireframe designs that can be expanded, rigidified or mechanically switched.:Abstract v Publications vii Acknowledgements ix Contents xiii Chapter 1 Introduction 1 1.1 Nanotechnology 1 1.1.1 History of nanotechnology 1 1.1.2 Phenomena that occur at nanoscale 4 1.1.3 Nature’s perspective of nanotechnology 4 1.1.4 Manufacturing nanomaterials 6 1.2 Deoxyribonucleic acid (DNA) 8 1.2.1 DNA, the genetic material, “The secret of life” 8 1.2.2 Structure of DNA 9 1.2.3 DNA synthesis 15 1.2.4 Stability of DNA 18 1.3 DNA nanotechnology 20 1.3.1 Historical development 20 1.3.2 DNA tile motifs 21 1.3.3 Directed nucleation assembly and algorithmic assembly 23 1.3.4 Scaffolded DNA origami and single-stranded DNA tiles 25 1.3.5 Expanding the design space offered by DNA 27 1.3.6 Assembling heterogeneous materials with DNA 30 1.3.7 Functional devices built using DNA nanostructures 35 Chapter 2 Motivation and objectives 40 Chapter 3 Block copolymer micellization as a protection strategy for DNA origami 42 3.1 Introduction 42 3.1.1 Cellular delivery of DNA nanostructures 42 3.1.2 The need for stability of DNA nanostructures 43 3.1.3 Non-viral gene therapy 44 3.2 Results and discussions 46 3.2.1 Strategy to form DNA origami polyplex micelles (DOPMs) 46 3.2.2 Optimizations 46 3.2.3 Decomplexation 53 3.2.4 Stability tests 55 3.2.5 Short PEG-PLys block copolymer 58 3.2.6 Compatibility with bulky ligands 59 3.2.7 Accessibility of handles on DOPMs 63 3.3 Conclusion 64 3.4 Outlook and state of the art 65 3.5 Methods 67 3.5.1 DNA origami folding 67 3.5.2 Preparation of ssDNA functionalized AuNPs 68 3.5.3 Agarose gel electrophoresis 69 3.5.4 Block copolymer preparation 70 3.5.5 DNA origami polyplex micelle preparation 70 3.5.6 Decomplexation of DOPM using dextran sulfate 73 3.5.7 Stability tests 74 3.5.8 tSEM characterization 75 3.5.9 AFM imaging 76 Chapter 4 Improving organic solubility and stability of DNA origami using polyplex micellization 77 4.1 Introduction 77 4.2 Results and discussions 79 4.2.1 Strategy for organic solubility of DNA origami 79 4.2.2 Proof of concept using AuNPs functionalized with ssDNA 80 4.2.3 Extending the strategy to DNA origami 82 4.2.4 Optimizations 86 4.2.5 Compatibility with functional ligands 88 4.2.6 Functionalization of DNA origami in organic solvent 94 4.3 Conclusion and outlook 95 4.4 Methods 97 4.4.1 Conjugation of functional ligands to DNA origami 97 4.4.2 Organic solubility 98 4.4.3 Reactions in organic solution on DOPMs 99 4.4.4 Fluorescence imaging using gel scanner 100 Chapter 5 Structural transformation of wireframe DNA origami via DNA polymerase assisted gap-filling 101 5.1 Introduction 101 5.2 Results and discussion 102 5.2.1 Design of the structures 102 5.2.2 Folding of gap-structures 105 5.2.3 Single-stranded DNA binding proteins 107 5.2.4 Gap filling with different polymerases 109 5.2.5 Gap filling with Phusion high-fidelity DNA polymerase 111 5.2.6 Optimization of the extension reaction using T4 DNA polymerase 115 5.2.7 Secondary structures 121 5.2.8 Folding kinetics of gap origami 124 5.2.9 Bending of tubes 125 5.3 Conclusion 126 5.4 Outlook 127 5.5 Methods 128 5.5.1 DNA origami folding 128 5.5.2 Gap filling of the wireframe DNA origami structures 128 5.5.3 Agarose gel electrophoresis 130 5.5.4 PAGE gel analysis 130 5.5.5 tSEM characterization 131 5.5.6 AFM imaging 131 5.5.7 AGE based folding-yield estimation 132 5.5.8 Gibbs free energy simulation using mfold 132 5.5.9 Staple list for folding the DNA origami triangulated structures 132 Appendix 134 A.1 Additional figures from chapter 3 134 A.2 Additional figures from chapter 4 137 A.3 Additional figures from chapter 5 149 Bibliography 155 Erklärung 171

Management of patients on long-term anticoagulation requiring percutaneous coronary intervention is challenging. Triple therapy with oral anticoagulant and dual antiplatelet therapy is the standard of care. However, there is no strong evidence to support this strategy. There is emerging data regarding the safety and efficacy of dual therapy with oral anticoagulant and single antiplatelet therapy in these patients. In this comprehensive review we highlight available evidence regarding various antithrombotic regimens’ efficacy and safety in patient with coronary artery disease undergoing percutaneous coronary intervention with long-term anticoagulation therapy requirements.