Strand Displacement Cascades Research Articles

G-Quadruplexes Light up Localized DNA Circuits.

DNA circuits tethered to nanoplatforms can perform cascade reactions for signal amplification. One DNA single strand activates a strand-displacement cascade generating numerous outputs, and therefore amplifying the signal. These localized circuits present, however, an important limitation: the spontaneous activation of the cascade reaction. Current methods to stabilize these circuits employ combination of protective DNA strands, which need to be removed to activate the device. This protection-deprotection process generates an important amount of unwanted side reactions. This is indeed an important limitation for the large potential application of these amplification circuits. In the present work, G-quadruplex DNA structures were used to stabilize localized DNA circuits. This new protocol generates nanoplatforms that no longer requires protective-deprotective systems and is therefore completely neutral to the sample. In addition, cations such as Pb(2+) or Ca(2+) can be also employed to activate the device enlarging the potential applications from biosensors devices to metal detector sensors.

A highly sensitive chemiluminescence sensor for detecting mercury (II) ions: a combination of Exonuclease III-aided signal amplification and graphene oxide-assisted background reduction

In this paper, we report a highly sensitive chemiluminescence (CL) sensor for Hg2+ ions based on thymine-Hg2+-thymine (T-Hg2+-T) coordination chemistry. We designed a thymine rich oligonucleotide as a capture probe and a signal probe that includes two functional domains: a horseradish peroxidase-mimicking DNAzyme domain for the generation of CL, and a recognition domain. Graphene oxide (GO) was introduced to adsorb the signal probe via p-p interaction, which brought the DNAzyme domain and GO into close proximity and quenches CL. In the presence of Hg2+ ions, the coordination of Hg2+ with the capture probe yielded a hairpin complex, triggers cascaded strand displacement reactions and Exonuclease III-assisted signal amplifications. As a result, accumulated amounts of DNAzyme were generated and released from GO, leading to an enhanced CL signal. This strategy combines enzyme-based signal amplification and GO as a background reducer, leads to a limit of detection (LOD) of 2 nmol/L. This simple detection system provides a label-free yet sensitive approach for detection of Hg2+ ions.

Real-time monitoring of enzyme-free strand displacement cascades by colorimetric assays.

The enzyme-free toehold-mediated strand displacement reaction has shown potential for building programmable DNA circuits, biosensors, molecular machines and chemical reaction networks. Here we report a simple colorimetric method using gold nanoparticles as signal generators for the real-time detection of the product of the strand displacement cascade. During the process the assembled gold nanoparticles can be separated, resulting in a color change of the solution. This assay can also be applied in complex mixtures, fetal bovine serum, and to detect single-base mismatches. These results suggest that this method could be of general utility to monitor more complex enzyme-free strand displacement reaction-based programmable systems or for further low-cost diagnostic applications.

On the biophysics and kinetics of toehold-mediated DNA strand displacement

Dynamic DNA nanotechnology often uses toehold-mediated strand displacement for controlling reaction kinetics. Although the dependence of strand displacement kinetics on toehold length has been experimentally characterized and phenomenologically modeled, detailed biophysical understanding has remained elusive. Here, we study strand displacement at multiple levels of detail, using an intuitive model of a random walk on a 1D energy landscape, a secondary structure kinetics model with single base-pair steps and a coarse-grained molecular model that incorporates 3D geometric and steric effects. Further, we experimentally investigate the thermodynamics of three-way branch migration. Two factors explain the dependence of strand displacement kinetics on toehold length: (i) the physical process by which a single step of branch migration occurs is significantly slower than the fraying of a single base pair and (ii) initiating branch migration incurs a thermodynamic penalty, not captured by state-of-the-art nearest neighbor models of DNA, due to the additional overhang it engenders at the junction. Our findings are consistent with previously measured or inferred rates for hybridization, fraying and branch migration, and they provide a biophysical explanation of strand displacement kinetics. Our work paves the way for accurate modeling of strand displacement cascades, which would facilitate the simulation and construction of more complex molecular systems.

Autonomous molecular cascades for evaluation of cell surfaces

Molecular automata are mixtures of molecules that undergo precisely defined structural changes in response to sequential interactions with inputs1–4. Previously studied nucleic acid-based-automata include game-playing molecular devices (MAYA automata3,5) and finite-state automata for analysis of nucleic acids6 with the latter inspiring circuits for the analysis of RNA species inside cells7,8. Here, we describe automata based on strand-displacement9,10 cascades directed by antibodies that can analyze cells by using their surface markers as inputs. The final output of a molecular automaton that successfully completes its analysis is the presence of a unique molecular tag on the cell surface of a specific subpopulation of lymphocytes within human blood cells.

DNA branch migration reactions through photocontrollable toehold formation.

Strand displacement cascades are commonly used to make dynamically assembled structures. Particularly, the concept of "toehold-mediated DNA branch migration reactions" has attracted considerable attention in relation to dynamic DNA nanostructures. However, it is a challenge to obtain and control the formation of pure 1:1 ratio DNA duplexes with toehold structures. Here, for the first time, we report a photocontrolled toehold formation method, which is based on the photocleavage of 2-nitrobenzyl linker-embedded DNA hairpin precursor structures. UV light irradiation (λ ≈ 365 nm) of solutions containing these DNA hairpin structures causes the complete cleavage of the nitrobenzyl linker, and pure 1:1 DNA duplexes with toehold structures are easily formed. Our experimental results indicate that the amount of toehold can be controlled by simply changing the dose of UV irradiation and that the resulting toehold structures can be used for subsequent toehold-mediated DNA branch migration reactions, e.g., DNA hybridization chain reactions. This newly established method will find broad application in the construction of light-powered, controllable, and dynamic DNA nanostructures or large-scale DNA circuits.

Neural network computation with DNA strand displacement cascades

The impressive capabilities of the mammalian brain--ranging from perception, pattern recognition and memory formation to decision making and motor activity control--have inspired their re-creation in a wide range of artificial intelligence systems for applications such as face recognition, anomaly detection, medical diagnosis and robotic vehicle control. Yet before neuron-based brains evolved, complex biomolecular circuits provided individual cells with the 'intelligent' behaviour required for survival. However, the study of how molecules can 'think' has not produced an equal variety of computational models and applications of artificial chemical systems. Although biomolecular systems have been hypothesized to carry out neural-network-like computations in vivo and the synthesis of artificial chemical analogues has been proposed theoretically, experimental work has so far fallen short of fully implementing even a single neuron. Here, building on the richness of DNA computing and strand displacement circuitry, we show how molecular systems can exhibit autonomous brain-like behaviours. Using a simple DNA gate architecture that allows experimental scale-up of multilayer digital circuits, we systematically transform arbitrary linear threshold circuits (an artificial neural network model) into DNA strand displacement cascades that function as small neural networks. Our approach even allows us to implement a Hopfield associative memory with four fully connected artificial neurons that, after training in silico, remembers four single-stranded DNA patterns and recalls the most similar one when presented with an incomplete pattern. Our results suggest that DNA strand displacement cascades could be used to endow autonomous chemical systems with the capability of recognizing patterns of molecular events, making decisions and responding to the environment.

Intercourse after Syphilis

Nucleic acid interactions under crowded environments are of great importance for biological processes and nanotechnology. However, the kinetics and thermodynamics of nucleic acid interactions in a crowded environment remain poorly understood. We use a coarse-grained model of DNA to study the kinetics and thermodynamics of DNA duplex and hairpin formation in crowded environments. We find that crowders can increase the melting temperature of both an 8-mer DNA duplex and a hairpin with a stem of 6-nt depending on the excluded volume fraction of crowders in solution and the crowder size. The crowding induced stability originates from the entropic effect caused by the crowding particles in the system. Additionally, we study the hybridization kinetics of DNA duplex formation and the formation of hairpin stems, finding that the reaction rate <i>k</i>_on is increased by the crowding effect, while <i>k</i>_off is changed only moderately. The increase in <i>k</i>_on mostly comes from increasing the probability of reaching a transition state with one base pair formed. A DNA strand displacement reaction in a crowded environment is also studied with the model and we find that rate of toehold association is increased, with possible applications to speeding up strand displacement cascades in nucleic acid nanotechnology.