We present a strategy for self-assembly of the smallest yet reported DNA nanostructures that are also addressable in terms of their DNA-base code. Using linear as well as novel branched threeway DNA oligonucleotide building-blocks we demonstrate the formation of a nano-network's fundamental cell, a DNA pseudo-hexagon of side 4 nm. The network's inherent addressability will allow functionalization with sub-nanometer precision yielding unprecedented richness in information density, important in context of Moore's Law and nano-chip technology.

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Here, we present the formation of a fully addressable DNA nanostructure that shows the potential to be exploited as, for example, an information storage device based on pH-driven triplex strand formation or nanoscale circuits based on electron transfer. The nanostructure is composed of two adjacent hexagonal unit cells (analogous to naphthalene) in which each of the eleven edges has a unique double-stranded DNA sequence, constructed using novel three-way oligonucleotides. This allows each ten base-pair side, just 3.4 nm in length, to be assigned a specific address according to its sequence. Such constructs are therefore an ideal precursor to a nonrepetitive two-dimensional grid on which the "addresses" are located at a precise and known position. Triplex recognition of these addresses could function as a simple yet efficient means of information storage and retrieval. Future applications that may be envisaged include nanoscale circuits as well as subnanometer precision in nanoparticle templating. Characterization of these precursor nanostructures and their reversible targeting by triplex strand formation is shown here using gel electrophoresis, atomic force microscopy, and fluorescence resonance energy transfer (FRET) measurements. The durability of the system to repeated cycling of pH switching is also confirmed by the FRET studies.

The chemical synthesis of oligonucleotides and their enzyme-mediated assembly into genes and genomes has significantly advanced multiple scientific disciplines. However, these approaches are not without their shortcomings; enzymatic amplification and ligation of oligonucleotides into genes and genomes makes automation challenging, and site-specific incorporation of epigenetic information and/or modified bases into large constructs is not feasible. Here we present a fully chemical one-pot method for the assembly of oligonucleotides into a gene by click-DNA ligation. We synthesize the 335 base-pair gene that encodes the green fluorescent protein iLOV from ten functionalized oligonucleotides that contain 5'-azide and 3'-alkyne units. The resulting click-linked iLOV gene contains eight triazoles at the sites of chemical ligation, and yet is fully biocompatible; it is replicated by DNA polymerases in vitro and encodes a functional iLOV protein in Escherichia coli. We demonstrate the power and potential of our one-pot gene-assembly method by preparing an epigenetically modified variant of the iLOV gene.