FOLDING DNA PERSPECTIVES fluorescent molecules, into well-defined geometries and stoichiometries, DNA origami enables precise control over nanoscale light – matter interactions. In this perspective, we discuss how DNA origami has been used to assemble nanophotonic systems and to study their interaction with single emitters.
DNA ORIGAMI FOR NANOPHOTONICS DNA ORIGAMI AS A BOTTOM-UP NANOFABRICATION TECHNIQUE In the early 1980s, Nadrian Seeman recognized that DNA could serve as a programmable material for the bottom-up fabrication of nanostructures. Singlestranded DNA( ssDNA) is a flexible polymer made of four nucleotides, adenine( A), thymine( T), guanine( G), and cytosine( C), with a persistence length of about 1 nm. When two strands carry complementary sequences, they can hybridize through Watson-Crick base pairing, A-T and G-C, into a double-stranded DNA( dsDNA) helix with a diameter of ~ 2 nm. This duplex is far more rigid, with a persistence length of approximately 50 nm. Introduced by Paul Rothemund in 2006, DNA origami exploits this predictable base-pairing rule to fold a long ssDNA scaffold into predefined twoor three-dimensional architectures with the help of hundreds of short synthetic“ staple” strands. [ 1 ]
Because each staple strand occupies a defined position within the origami design, the resulting structure acts as an addressable molecular breadboard. Selected staples can be chemically modified, for example with fluorophores, or extended with short single-stranded DNA handles that serve as programmable docking sites. NPs, quantum dots, or other optical components functionalized with the complementary DNA sequence can then be selectively attached to these positions( Figure 2). In this way, DNA origami enables the construction of hybrid photonic structures in which the nature, number, and relative spacing of the components are defined at the sequence level.
This bottom-up strategy addresses several limitations of conventional topdown nanofabrication. First, DNA origami relies on commercially available
THE DNA ORIGAMI WORKFLOW: FROM DESIGN TO SELF-ASSEMBLY
1. Digital design( caDNAno): The process starts from the target geometry, which is defined according to the optical function to be implemented and the required spatial arrangement of the components. The DNA origami host is then designed with computer-aided( CAD) design tools such as caDNAno, which allow the scaffold strand( with around 7-8 thousand nucleotides) to be routed through a set of parallel DNA helices arranged, for example, on a square or honeycomb lattice. The choice of scaffold, often derived from the M13 bacteriophage genome, fixes the available contour length and therefore constrains the final size of the structure, typically from several tens to a few hundred nanometers.
2. Staple generation: Once the scaffold path has been fixed, the design software generates the sequences of short synthetic DNA oligonucleotides( i. e., the staple strands). These staples bind selected regions of the scaffold and force it to fold into the prescribed shape. A typical origami structure contains a few hundred staples, although the exact number depends on the scaffold length and the complexity of the design. Several commercial vendors specialize in automated DNA synthesis of ssDNA.
3. Thermal annealing: The scaffold and staple strands are mixed in buffer, generally with Mg 2 + ions to screen the electrostatic repulsion between DNA backbones. In practice, each staple strand is added in molar excess relative to the scaffold, typically at a 5- to 10-fold excess, to favor complete hybridization of the scaffold during folding. The mixture is heated and then slowly cooled, allowing the staples to bind their target scaffold regions and drive the folding process. Depending on the structure, annealing can take from a few hours to several days. Excess, non-incorporated staples are then removed, for example by gel electrophoresis or centrifugal filtration.
4. Structural characterization: The folded nanostructures are finally checked by imaging methods such as atomic force microscopy( AFM) or transmission electron microscopy( TEM). These measurements verify the folding yield and structural integrity.
Figure 1. Computer-aided design of DNA origami. DNA origami structures are usually designed with the caDNAno software, that can generate the sequences of the“ staple” strands required for the assembly of the designed geometry. Those staple strands are mixed with the“ scaffold” with a large excess to assemble the origami structures through thermal annealing. Proper folding is verified with different techniques. Figure adapted from [ 2 ].
scaffolds and staple strands, and can be implemented in standard chemistry or biochemistry laboratories without the need for clean-room facilities. Moreover, self-assembly is intrinsically parallel, allowing a single reaction to produce very large numbers of nominally identical nanostructures. Second, the method can incorporate colloidal metallic NPs, which are available in a wide
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