Facile and Scalable Preparation of Pure and Dense DNA Origami Solutions**

DNA has become a prime material for assembling complex three-dimensional objects that promise utility in various areas of application. However, achieving user-defined goals with DNA objects has been hampered by the difficulty to prepare them at arbitrary concentrations and in user-defined solution conditions. Here, we describe a method that solves this problem. The method is based on poly(ethylene glycol)-induced depletion of species with high molecular weight. We demonstrate that our method is applicable to a wide spectrum of DNA shapes and that it achieves excellent recovery yields of target objects up to 97 %, while providing efficient separation from non-integrated DNA strands. DNA objects may be prepared at concentrations up to the limit of solubility, including the possibility for bringing DNA objects into a solid phase. Due to the fidelity and simplicity of our method we anticipate that it will help to catalyze the development of new types of applications that use self-assembled DNA objects.

[1]  H. Dietz,et al.  Quantifying quality in DNA self-assembly , 2014, Nature Communications.

[2]  William M. Shih,et al.  Virus-Inspired Membrane Encapsulation of DNA Nanostructures To Achieve In Vivo Stability , 2014, ACS nano.

[3]  V. Linko,et al.  The enabled state of DNA nanotechnology. , 2013, Current opinion in biotechnology.

[4]  M. Rief,et al.  Rigid DNA Beams for High-Resolution Single-Molecule Mechanics** , 2013, Angewandte Chemie.

[5]  William M Shih,et al.  DNA nanotubes for NMR structure determination of membrane proteins , 2013, Nature Protocols.

[6]  Chenxiang Lin,et al.  Purification of DNA-origami nanostructures by rate-zonal centrifugation , 2012, Nucleic acids research.

[7]  T. G. Martin,et al.  Rapid Folding of DNA into Nanoscale Shapes at Constant Temperature , 2012, Science.

[8]  Luvena L. Ong,et al.  Three-Dimensional Structures Self-Assembled from DNA Bricks , 2012, Science.

[9]  T. G. Martin,et al.  Cryo-EM structure of a 3D DNA-origami object , 2012, Proceedings of the National Academy of Sciences.

[10]  T. G. Martin,et al.  Synthetic Lipid Membrane Channels Formed by Designed DNA Nanostructures , 2012, Science.

[11]  Samara L. Reck-Peterson,et al.  Tug-of-War in Motor Protein Ensembles Revealed with a Programmable DNA Origami Scaffold , 2012, Science.

[12]  Hendrik Dietz,et al.  Magnesium-free self-assembly of multi-layer DNA objects , 2012, Nature Communications.

[13]  Yonggang Ke,et al.  Two design strategies for enhancement of multilayer-DNA-origami folding: underwinding for specific intercalator rescue and staple-break positioning. , 2012, Chemical science.

[14]  P. Yin,et al.  Complex shapes self-assembled from single-stranded DNA tiles , 2012, Nature.

[15]  T. G. Martin,et al.  DNA origami gatekeepers for solid-state nanopores. , 2012, Angewandte Chemie.

[16]  Shawn M. Douglas,et al.  A Logic-Gated Nanorobot for Targeted Transport of Molecular Payloads , 2012, Science.

[17]  M. Hagan,et al.  Reconfigurable self-assembly through chiral control of interfacial tension , 2012, Nature.

[18]  K. Gothelf,et al.  Multilayer DNA origami packed on hexagonal and hybrid lattices. , 2012, Journal of the American Chemical Society.

[19]  M. Bathe,et al.  Quantitative prediction of 3D solution shape and flexibility of nucleic acid nanostructures , 2011, Nucleic acids research.

[20]  F. Simmel,et al.  DNA-based self-assembly of chiral plasmonic nanostructures with tailored optical response , 2011, Nature.

[21]  Hao Yan,et al.  Challenges and opportunities for structural DNA nanotechnology. , 2011, Nature nanotechnology.

[22]  S. Harrison,et al.  Mitochondrial uncoupling protein 2 structure determined by NMR molecular fragment searching , 2011, Nature.

[23]  Hao Yan,et al.  DNA Origami with Complex Curvatures in Three-Dimensional Space , 2011, Science.

[24]  Mark Bathe,et al.  A primer to scaffolded DNA origami , 2011, Nature Methods.

[25]  Chenxiang Lin,et al.  Recovery of intact DNA nanostructures after agarose gel–based separation , 2011, Nature Methods.

[26]  A. Turberfield,et al.  DNA-templated protein arrays for single-molecule imaging. , 2011, Nano letters.

[27]  N. Seeman Nanomaterials based on DNA. , 2010, Annual review of biochemistry.

[28]  Friedrich C. Simmel,et al.  DNA Origami as a Nanoscopic Ruler for Super‐Resolution Microscopy , 2009 .

[29]  Philip Tinnefeld,et al.  DNA‐Origami als Nanometerlineal für die superauflösende Mikroskopie , 2009 .

[30]  Shawn M. Douglas,et al.  Multilayer DNA origami packed on a square lattice. , 2009, Journal of the American Chemical Society.

[31]  Shawn M. Douglas,et al.  Folding DNA into Twisted and Curved Nanoscale Shapes , 2009, Science.

[32]  Viktor Stein,et al.  An efficient method to assemble linear DNA templates for in vitro screening and selection systems , 2009, Nucleic acids research.

[33]  Adam H. Marblestone,et al.  Rapid prototyping of 3D DNA-origami shapes with caDNAno , 2009, Nucleic acids research.

[34]  Elodie Boisselier,et al.  Gold nanoparticles in nanomedicine: preparations, imaging, diagnostics, therapies and toxicity. , 2009, Chemical Society reviews.

[35]  J. Kjems,et al.  Self-assembly of a nanoscale DNA box with a controllable lid , 2009, Nature.

[36]  Shawn M. Douglas,et al.  Self-assembly of DNA into nanoscale three-dimensional shapes , 2009, Nature.

[37]  B. Vajda Concentration and purification of viruses and bacteriophages with polyethylene glycol , 2008, Folia Microbiologica.

[38]  José María Carazo,et al.  Image processing for electron microscopy single-particle analysis using XMIPP , 2008, Nature Protocols.

[39]  Shawn M. Douglas,et al.  DNA-nanotube-induced alignment of membrane proteins for NMR structure determination , 2007, Proceedings of the National Academy of Sciences.

[40]  Wen Jiang,et al.  EMAN2: an extensible image processing suite for electron microscopy. , 2007, Journal of structural biology.

[41]  P. Rothemund Folding DNA to create nanoscale shapes and patterns , 2006, Nature.

[42]  Xie Hong-kun,et al.  Nature of Science , 2002 .

[43]  A. Minton,et al.  Macromolecular crowding: biochemical, biophysical, and physiological consequences. , 1993, Annual review of biophysics and biomolecular structure.

[44]  J. Sambrook,et al.  Molecular Cloning: A Laboratory Manual , 2001 .

[45]  P. Borst,et al.  A freeze-squeeze method for recovering long DNA from agarose gels. , 1975, Analytical biochemistry.

[46]  E. S. Anderson,et al.  A simple method for the preparation of large quantities of pure plasmid DNA. , 1975, Biochimica et biophysica acta.

[47]  B. Alberts,et al.  Rapid bacteriophage sedimentation in the presence of polyethylene glycol and its application to large-scale virus purification. , 1970, Virology.