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Abstract DNA‐based computers can potentially analyze complex sets of biological markers, thereby advancing diagnostics and the treatment of diseases. Despite extensive efforts, DNA processors have not yet been developed due, in part, to limitations in the ability to integrate available logic gates into circuits. We have designed a NAND gate, which is one of the functionally complete set of logic connectives. The gate's design avoids stem‐loop‐folded DNA fragments, and is capable of reusable operations in RNase H‐containing buffer. The output of the gate can be translated into RNA‐cleaving activity or a fluorescent signal produced either by a deoxyribozyme or a molecular beacon probe. Furthermore, three NAND‐gate‐forming DNA strands were crosslinked by click chemistry and purified in a simple procedure that allowed ≈10¹³gates to be manufactured in 16 h, with a hands‐on time of about 30 min. Two NAND gates can be joined into one association that performs a new logic function simply by adding a DNA linker strand. Approaches developed in this work could contribute to the development of biocompatible DNA logic circuits for biotechnological and medical applications. 

Accessibility of synthetic oligonucleotides and the success of DNA nanotechnology open a possibility to use DNA nanostructures for building sophisticated enzyme-like catalytic centers. Here we used a double DNA crossover (DX) tile nanostructure to enhance the rate, the yield, and the specificity of 5′−5′ ligation of two oligonucleotides with arbitrary sequences. The ligation product was isolated via a simple procedure. The same strategy was applied for the synthesis of 3′−3′ linked oligonucleotides, thus introducing a synthetic route to DNA and RNA with a switched orientation that is affordable by a low- resource laboratory. To emphasize the utility of the ligation products, we synthesized a circular structure formed from intramolecular complementarity that we named “an impossible DNA wheel” since it cannot be built from regular DNA strands by enzymatic reactions. Therefore, DX-tile nanostructures can open a route to producing useful chemical products that are unattainable via enzymatic synthesis. This is the first example of the use of DNA nanostructures as a catalyst. This study advocates for further exploration of DNA nanotechnology for building enzyme-like reactive systems. 

Hybridization probes have been used in the detection of specific nucleic acids for the last 50 years. Despite the extensive efforts and the great significance, the challenges of the commonly used probes include (1) low selectivity in detecting single nucleotide variations (SNV) at low (e.g. room or 37 °C) temp- eratures; (2) low affinity in binding folded nucleic acids, and (3) the cost of fluorescent probes. Here we introduce a multicomponent hybridization probe, called OWL2 sensor, which addresses all three issues. The OWL2 sensor uses two analyte binding arms to tightly bind and unwind folded analytes, and two sequence-specific strands that bind both the analyte and a universal molecular beacon (UMB) probe to form fluorescent ‘OWL’ structure. The OWL2 sensor was able to differentiate single base mismatches in folded analytes in the temperature range of 5–38 °C. The design is cost-efficient since the same UMB probe can be used for detecting any analyte sequence. 

Multiplex assays often rely on expensive sensors incorporating covalently linked fluorescent dyes. Herein, we developed a self-assembling aptamer-based multiplex assay. This multiplex approach utilizes a previously established split aptamer sensor in conjugation with a novel split aptamer sensor based upon a malachite green DNA aptamer. This system was capable of simultaneous fluorescent detection of two SARS COVID-19-related sequences in one sample with individual sensors that possesses a limit of detection (LOD) in the low nM range. Optimization of the Split Malachite Green (SMG) sensor yielded a minimized aptamer construct, Mini-MG, capable of inducing fluorescence of malachite green in both a DNA hairpin and sensor format. 

DNA nanotechnology uses oligonucleotide strands to assemble molecular structures capable of performing useful operations. Here, we assembled a multifunctional prototype DNA nanodevice, DOCTR, that recognizes a single nucleotide mutation in a cancer marker RNA. The nanodevice then cuts out a signature sequence and uses it as an activator for a "therapeutic" function, namely, the cleavage of another RNA sequence. The proposed design is a prototype for a gene therapy DNA machine that cleaves a housekeeping gene only in the presence of a cancer-causing point mutation and suppresses cancer cells exclusively with minimal side effects to normal cells.