DNA nanotechnology | materials and methods

Materials and methods

Gel electrophoresis methods, such as this formation assay on a DX complex, are used to ascertain whether the desired structures are forming properly. Each vertical lane contains a series of bands, where each band is characteristic of a particular reaction intermediate.

The sequences of the DNA strands making up a target structure are designed computationally, using molecular modeling and thermodynamic modeling software. [67] [69] The nucleic acids themselves are then synthesized using standard oligonucleotide synthesis methods, usually automated in an oligonucleotide synthesizer, and strands of custom sequences are commercially available. [70] Strands can be purified by denaturing gel electrophoresis if needed, [71] and precise concentrations determined via any of several nucleic acid quantitation methods using ultraviolet absorbance spectroscopy. [72]

The fully formed target structures can be verified using native gel electrophoresis, which gives size and shape information for the nucleic acid complexes. An electrophoretic mobility shift assay can assess whether a structure incorporates all desired strands. [73] Fluorescent labeling and Förster resonance energy transfer (FRET) are sometimes used to characterize the structure of the complexes. [74]

Nucleic acid structures can be directly imaged by atomic force microscopy, which is well suited to extended two-dimensional structures, but less useful for discrete three-dimensional structures because of the microscope tip's interaction with the fragile nucleic acid structure; transmission electron microscopy and cryo-electron microscopy are often used in this case. Extended three-dimensional lattices are analyzed by X-ray crystallography. [75] [76]

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