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Background: Differentiating between different states in nucleic acid circuits is crucial for various biological applications. One approach, there is a requirement for complicated sequential summation, which can be excessive for practical purposes. By selectively labeling biologically significant states, this study tackles the issue and presents a more cost-effective and streamlined solution. The challenge is to efficiently distinguish between different states in a nucleic acid circuit., Results: An innovative method is introduced in this study to distinguish between states in a nucleic acid circuit, emphasizing the biologically relevant ones. The circuit comprises four DNA logic gates and two detection modules, one for determining fetal gender and the other for diagnosing X-linked genetic disorders. The primary module generates a G-quadruplex DNAzyme when activated by specific biomarkers, which leads to a distinct colorimetric signal. The secondary module responds to hemophilia and choroideremia biomarkers, generating one or two DNAzymes. The absence of female fetus indicators results in no DNAzyme or color change. The circuit can differentiate various fetal states by producing one to four active DNAzymes in response to male fetus biomarkers. A single-color solution for state differentiation is provided by this approach, which promises significant advancements in DNA computing and diagnostic applications., Significance: The innovative approach used in this study to distinguish states in nucleic acid circuits holds great significance. By selectively labeling biologically relevant states, circuit design is simplified and complexity is reduced. This advancement enables cost-effective and efficient diagnostic applications and contributes to DNA computing, providing a valuable solution to a fundamental problem., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Developing isothermal bio-analyzers for amplified detection of multi-factor diseases like cancer biomarkers (nucleic acid and protein) has facilitated the early diagnosis and clinical theranostics. In light of that, a sensitive detection system was developed assisted by the recognition capability of a functional aptamer followed by cyclic self-assembly of three auxiliary hairpins via branched hybridization chain reaction (b-HCR) performance. In the downstream process, in the presence of hemin, split sequences of a DNAzyme brought in close proximity to facilitate the color alteration of the solution to a green appearance. By ingenious exerting multi-level amplification, the assay empowered sensitive detection of miR-21 and PDGF-BB related cancer biomarkers with LOD values of 10 pM and 40 nM, respectively. Taken together, simplicity, enzyme-free nature, excellent generality, affordable cost without any washing steps and immobilization makes the presented system a promising analytical tool in synthetic biology, designing nanomachines and point-of-care detection in resource-constrained settings., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022. Published by Elsevier B.V.)

The development of powerful techniques for sensitive detection of nucleic acids has attracted much attention for fabricating accurate biosensors in various fields, such as genomics, clinical diagnostics, and forensic sciences. Up to now, different systems have been introduced, the majority of which are expensive, time-consuming, and relatively low selectivity/limit of detection. These limitations caught our attention to fabricate a nucleic acid responsive system by combining three layers of signal amplification strategy, namely a split proximity circuit (SPC), a catalytic hairpin assembly (CHA), and a DNA hydrogel. Herein, by SPC operation, two initiators and a target strand were assembled and activated the CHA reaction in the presence of three 5'-cytosine (C)-rich hairpins. Then, produced C-rich embedded three-way junction structures could form i-motif structures under acidic environment followed by a transition from sol to gel state. To acquire a quantitative and colorimetric measurement, gold nanoparticles (GNPs) were used that encapsulated and sediment by the gel formation. The resulting platform detected the target with a limit of detection of 1 pM and considerable selectivity., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Fluctuation of nucleic acid expression and ultrasensitive and specific detection of these variations in expression is a crucial subject in molecular medicine and clinical theranostics. A novel DNAzyme-coupled branched hybridization chain reaction (b-HCR) assay is reported for efficient signal-amplified detection of miRNA in this study. This assay was composed of a translator (T) hybridized with miR-21 to initiate the first HCR by hairpin 1 (H 1 ) and hairpin 2 (H 2 ). The primary HCR provided a backbone chain for numerous branches budding through hairpin 3 (H 3 ) and hairpin 4 (H 4 ) assembles. In the presence of hemin, the G-rich domains embedded in H1 and H4 produce an active G-quadruplex DNAzyme upon exposure to a target that could catalyze the oxidation of colorless substrate to colored product. The present approach has the potential to be used for quantitative detection of miR-21 with a sensitivity and a dynamic range of 1 pM and 1 pM to 1 nM, respectively., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020. Published by Elsevier B.V.)

An enzyme-free dual catalytic DNA circuit for amplified detection of nucleic acids has been developed. The system functions based on a cyclic self-assembly of two auxiliary hairpins (H1 and H2) and three biotinylated hairpin oligonucleotides (H3, H4 and H5), in the format of two molecular circuits. In the upstream circuit, a target initiator (I) besides H1 and H2 hairpins constructs H1-H2 duplexes that trigger the operation of a subsequent circuit. In the downstream circuit, the H1-H2 duplex initiates cascaded self-assembly reactions, produces triplex H3-H4-H5, as sensing system, and releases the H1-H2 duplex as the catalyst for the self-assembly of additional hairpins. The H3-H4-H5 triplex acts as the scaffolds for assembling and orienting the streptavidin-functionalized gold nanoparticles (SA-AuNPs) into a lattice-like arrangement that generates a DNA-SA-AuNP cross-linked network, resulting in a dramatic pale red-to-blue color change. By ingeniously engaging two catalytic circuits with feedback amplification capabilities, the system can detect the target nucleic acid with an LOD value of 5 femtomolar and unambiguously discriminate spurious targets (i.e. targets containing substitution, insertion, and deletion nucleotides) without instrumentation. Simple and convenient operation of the assay makes the DNA circuit appropriate for point-of-care monitoring in resource-constrained settings., (Copyright © 2020. Published by Elsevier B.V.)

A DNAzyme-embedded hyperbranched DNA dendrimer is used as a colorimetric signal amplifier in an ultrasensitive detection scheme for nucleic acids. The hyperbranched DNA dendrimers were constructed by single-step autonomous self-assembly of three structure-free DNA monomers. A cascade of self-assembly reactions between the first and second strands leads to the formation of linear DNA concatemers containing overhang flank fragments. The third strand, which bears a peroxidase-mimicking DNAzyme domain, serves as a bridge to trigger self-assembly between the first and second strands across the side chain direction. This results in a chain branching growth of the DNAzyme-embedded DNA dendrimer. This signal amplifier was incorporated into the streptavidin-biotin detection system which comprises an adaptor oligonucleotide and a biotinylated capture probe. The resulting platform is capable of detecting a nucleic acid target with an LOD as low as 0.8 fM. Such sensitivity is comparable if not superior to most of the reported enzyme-free (and even enzyme-assisted) signal amplification strategies. The DNA dendrimer based method is expected to provide a universal platform for extraordinary signal enhancement in detecting other nucleic acid biomarkers by altering the respective sequences of adaptor and capture probe. Graphical abstract Schematic of an assembly of a DNAzyme-embedded hyperbranched DNA dendrimer which operates as a signal amplifier for nucleic acids detection. The nanostructure is constructed by autonomous self-assembly of three DNA monomers. Colored letters represent each domain, and complementary domains are marked by asterisk. Domain d represents the DNAzyme sequence.

In the present study, a highly sensitive and specific bio-sensing platform for enzyme-free and colorimetric detection of nucleic acids has been developed. The biosensor is composed of two DNA nanostructures and two fuel strands that construct the foundation of a feed-forward catalytic DNA circuit. Upon binding the target strand to a specific DNA nanostructure, the circuit is run in order that at the end a hemin-binding aptamer, with the ability to convert a colorless substrate into a colored substance is released. Based on this strategy, 4 pM of the target DNA can be easily detected in serum samples by naked eyes after only a two-hour incubation with the circuit; meanwhile, if the incubation time is extended to 3 h, the biosensor can detect 1 pM of the target DNA. Besides the elevated sensitivity, the circuit can truly discriminate a spurious target containing one nucleotide mismatch with high specificity. Overall, the enzyme-free catalytic DNA circuit can be used as a sensitive alternative method to enzyme-based biosensors for the specific and cost-effective detection of nucleic acids., (Copyright © 2016 Elsevier B.V. All rights reserved.)