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t has been shown that active learning strategies are effective in teaching complex STEM concepts. In this study, we developed and implemented a laboratory experiment for teaching the concepts of Boolean logic gates, molecular beacon probes, molecular computing, DNA logic gates, microRNA, and molecular diagnosis of hepatocellular carcinoma, which are related to DNA molecular computing, an interdisciplinary cutting-edge research technology in biochemistry, synthetic biology, computer science, and medicine. The laboratory experience takes about 110–140 min and consists of a multiple-choice pretest (15 min), introductory lecture (20 min), wet laboratory experiment (60–90 min), and a post-test (15 min). Students are tasked to experimentally construct three molecular logic circuits made of DNA oligonucleotides and use them for the fluorescence-based detection of microRNA markers related to diagnostics of hepatocellular carcinoma. The class was taught to undergraduate students from freshman to senior academic levels majoring in chemistry, biochemistry, biotechnology, and biomedical sciences. Students were engaged during the session and motivated to learn more about the research technology. A comparison of students’ scores on the pretest and post-test demonstrated improvement in knowledge of the concepts taught. Visual observation of the fluorescence readout led to a straightforward interpretation of the results. The laboratory experiment is portable; it uses inexpensive nontoxic reagents and thus can be employed outside a laboratory room for outreach and science popularization purposes. 

A functionally complete Boolean operator is sufficient for computational circuits of arbitrary complexity. We connected YES (buffer) with NOT (inverter) and two NOT four-way junction (4J) DNA gates to obtain IMPLY and NAND Boolean functions, respectively, each of which represents a functionally complete gate. The results show a technological path towards creating a DNA computational circuit of arbitrary complexity based on singleton NOT or a combination of NOT and YES gates, which is not possible in electronic computers. We, therefore, concluded that DNA-based circuits and molecular computation may offer opportunities unforeseen in electronics. 

Due to nucleic acid's programmability, it is possible to realize DNA structures with computing functions, and thus a new generation of molecular computers is evolving to solve biological and medical problems. Pioneered by Milan Stojanovic, Boolean DNA logic gates created the foundation for the development of DNA computers. Similar to electronic computers, the field is evolving towards integrating DNA logic gates and circuits by positioning them on substrates to increase circuit density and minimize gate distance and undesired crosstalk. In this minireview, we summarize recent developments in the integration of DNA logic gates into circuits localized on DNA substrates. This approach of all‐DNA integrated circuits (DNA ICs) offers the advantages of biocompatibility, increased circuit response, increased circuit density, reduced unit concentration, facilitated circuit isolation, and facilitated cell uptake. DNA ICs can face similar challenges as their equivalent circuits operating in bulk solution (bulk circuits), and new physical challenges inherent in spatial localization. We discuss possible avenues to overcome these obstacles. 

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. 

Molecular beacon (MB) probes have been extensively used for nucleic acid analysis. However, MB probes fail to hybridize with folded DNA or RNA. Here, we demonstrate that MB probes equipped with extra sequences complementary to the analyte, named ‘tail’, can increase the signal-to-background ratio by B40- fold and hybridization rates by B800-fold compared to conventional MB probes. Tailed MB probes can be used as mismatched-tolerant alternatives to traditional hairpin probes for fast assays. 

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) temperatures; (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. 

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.