Stability of DNA Origami Nanoarrays in Cell Lysate

Abstract

Scaffolded DNA origami, a method to create self-assembled nanostructures with spatially addressable features, has recently been used to develop water-soluble molecular chips for label-free RNA detection, platforms for deterministic protein positioning, and single molecule reaction observatories. These applications highlight the possibility of exploiting the unique properties and biocompatibility of DNA nanostructures in live, cellular systems. Herein, we assembled several DNA origami nanostructures of differing shape, size and probes, and investigated their interaction with lysate obtained from various normal and cancerous cell lines. We separated and analyzed the origami–lysate mixtures using agarose gel electrophoresis and recovered the DNA structures for functional assay and subsequent microscopic examination. Our results demonstrate that DNA origami nanostructures are stable in cell lysate and can be easily separated from lysate mixtures, in contrast to natural, single- and double-stranded DNA. Atomic force microscope (AFM) and transmission electron microscope (TEM) images show that the DNA origami structures are fully intact after separation from cell lysates and hybridize to their targets, verifying the superior structural integrity and functionality of self-assembled DNA origami nanostructures relative to conventional oligonucleotides. The stability and functionality of DNA origami structures in cell lysate validate their use for biological applications, for example, as programmable molecular rafts or disease detection platforms.

DNA Nanostructures as Smart Drug-Delivery Vehicles and Molecular Devices

“…One of them is to improve the pharmacokinetic bioavailability of the DNA-based structures in vivo. DNA origami structures can survive in cellular milieu [22] and against nucleases [20], but the circulation times of the objects could be increased by adjusting their modular properties (see above) or by creating novel protecting strategies such as protein- [47] or lipid membrane coatings [48].Another challenge in creating DNA-based delivery vehicles and nanomachines is arguably the relatively expensive synthesis of the starting materials. However, since the entire research field is constantly growing and developing, it is likely that scaling up of the production and simultaneous knockdown of the price will take place rapidly [3,64].…”

Temperature-Controlled Encapsulation and Release of an Active Enzyme in the Cavity of a Self-Assembled DNA Nanocage

“…34 More recently DNA origami nanoarrays were demonstrated stable in cell lysates. 35 DNA cage structures have also been demonstrated capable of entering live mammalian cells, 36 while DNA tubes or barrels have been demonstrated directable to specific cell types to which they can either deliver 37 or expose 31 their specific cargo.…”

Drug delivery systems based on nucleic acid nanostructures

“…Before the DNA origami structures were introduced for biomedical purposes their stability in cell lysates was investigated [77]. Four assemblies were fabricated for that purpose: a 2D rectangular origami (90 × 60 nm) [10], a 2D equilateral triangle (120 nm long with 30 nm wide sides) with an open central triangular cavity of 60 nm per side, a 3D multilayer rectangular parallelepiped structure (16 × 16 × 30 nm) and a 2D rectangular structure with staple strands bearing overhangs allowing hybridization (Fig.…”

“…11). It is based on an 8634 nt long scaffold and during hybridization of staples every seventh nt an insertion was introduced similar as reported before [80] Reproduced with permission from [77] and [10].…”