Programming the Sequential Release of DNA

Scalise, Dominic; Rubanov, Moshe; Miller, Katherine; Potters, Leo; Noble, Madeline; Schulman, Rebecca

doi:10.1021/acssynbio.9b00398

Cited by 24 publications

(45 citation statements)

References 36 publications

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“…2C). We opted to use a DNA-based reporter, rather than an RNA aptamerbased reporter (46,47), because the DNA reporter is easily calibrated to output concentration for modeling (4,5,7). To be stable at 37 o C, we designed the reporter with a 16 base duplex.…”

Section: Experimental Characterization and Modeling Of Ctrsdmentioning

confidence: 99%

“…To develop ctRSD, we sought a system in which modular and programmable strand displacement circuit components could be isothermally produced via transcription. In TMSD circuits, modularity is achieved by designing toehold exchange gates that allow any input sequence to be converted into any output sequence through a gate (4,5,7,12). For example, in Fig.…”

Section: Design Of Co-transcriptional Rna Strand Displacement (Ctrsd) Componentsmentioning

confidence: 99%

“…We restricted the gate output sequences to cytosine (C), adenine (A), or uracil (U) bases. This sequence constraint reduces unwanted secondary structure or dimerization of single-stranded components (4,5,7). A G-U wobble pair was also introduced in the middle of the hybridized portion of the gate to reduce DNA template synthesis errors (40) and to drive the forward ctRSD reaction with inputs that convert the G-U wobble to a G-C pair (41).…”

Section: Design Of Co-transcriptional Rna Strand Displacement (Ctrsd) Componentsmentioning

confidence: 99%

“…We next investigated whether ctRSD components could be programed to execute logic (5,48), signal amplification (6,8), and multi-layer cascades (7). To assess the predictability of ctRSD circuit design, for each circuit we built we evaluated how well our kinetic model predicted behavior.…”

Section: Ctrsd Logic and Signal Amplification Elementsmentioning

confidence: 99%

“…The predictable and programmable Watson-Crick base pairing interactions of nucleic acids has led to their adoption as versatile components for molecular circuit programming. In particular, in vitro circuits based on toehold-mediated strand displacement (TMSD) reactions have demonstrated sophisticated digital computations and mathematical operations (5), molecular pattern recognition (4,6), signal cascades (7) and amplifiers (5,8,9), and complex dynamics (9)(10)(11). In TMSD reactions, a single-stranded input binds to a double-stranded nucleic acid gate via a single-stranded toehold domain and displaces an output strand with a new exposed toehold that can facilitate further TMSD reactions (Fig.…”

Section: Introductionmentioning

confidence: 99%

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Co-transcriptional RNA strand displacement circuits

Schaffter

Strychalski

2021

Preprint

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Engineered molecular circuits that process information in biological systems could address emerging human health and biomanufacturing needs. However, such circuits can be difficult to rationally design and scale. DNA-based strand displacement reactions have demonstrated the largest and most computationally powerful molecular circuits to date but are limited in biological systems due to the difficulty in genetically encoding components. Here, we develop scalable co-transcriptional RNA strand displacement (ctRSD) circuits that are rationally programmed via base pairing interactions. ctRSD addresses the limitations of DNA-based strand displacement circuits by isothermally producing circuit components via transcription. We demonstrate the programmability of ctRSD in vitro by implementing logic and amplification elements, and multi-layer signaling cascades. Further, we show ctRSD kinetics are accurately predicted by a simple model of coupled transcription and strand displacement, enabling model-driven design. We envision ctRSD will enable rational design of powerful molecular circuits that operate in biological systems, including living cells.

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Section: Experimental Characterization and Modeling Of Ctrsdmentioning

confidence: 99%

Section: Design Of Co-transcriptional Rna Strand Displacement (Ctrsd) Componentsmentioning

confidence: 99%

Section: Design Of Co-transcriptional Rna Strand Displacement (Ctrsd) Componentsmentioning

confidence: 99%

Section: Ctrsd Logic and Signal Amplification Elementsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Co-transcriptional RNA strand displacement circuits

Schaffter

Strychalski

2021

Preprint

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Engineering Aptamer Switches for Multifunctional Stimulus‐Responsive Nanosystems

et al. 2020

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specific and strong recognition of molecular targets (i.e., aptamers), catalyze biochemical reactions (i.e., ribozymes), or form sophisticated and dynamic "smart materials" or even 3D constructs (i.e., DNA origami). [2] There is particular interest in the use of nucleic acids for the construction of molecular switches, which undergo a function-altering structural change in response to an external stimulus. [3] Many such examples exist in nature, where switches enable organisms to sense physiological changes and selectively control biological functions in response. For example, riboswitches exist naturally as domains within messenger RNA (mRNA) transcripts that can bind to specific metabolites with high specificity in order to stabilize a secondary structure that prevents subsequent translation. [4] Synthetic, engineered molecular switches based on nucleic acids can achieve even greater diversity of function. Such designs make use of aptamersnucleic acid-based affinity reagents isolated via an in vitro process of systematic evolution by exponential enrichment (SELEX), in which DNA or RNA sequences with specific ligand binding are isolated from a large pool of random sequences. [5] Aptamers have proven to be robust recognition elements due to their thermal stability, ease of synthesis, and capacity to undergo ready modification with a broad array of functional groups. Importantly, traditional selection methods can be expanded upon by innovative engineering techniques to enable the incorporation of additional functions that further extend the utility of aptamers. Aptamers can adopt various 2-and 3-D structures such as duplexes, hairpins, and G-quadruplexes, and the inducible formation and disruption of these configurations can be exploited to enable complex functions besides simple biosensing. Consequently, aptamers have been incorporated into many intricate configurations, such as logic gates and dynamic nanomachines. Various research groups have also described aptamer-based switches that undergo conformational changes in response to triggers such as shifts in pH, stimulation with light, or the presence of a specific ligand. [6] Although still a relatively young area of research, these efforts could prove highly useful for materials research-conferring even greater and more selective control over devices based on functionalized nucleic acids. Aptamers are becoming increasingly integrated with various organic and inorganic molecules in order to control the assembly and operation of novel functional Although RNA and DNA are best known for their capacity to encode biological information, it has become increasingly clear over the past few decades that these biomolecules are also capable of performing other complex functions, such as molecular recognition (e.g., aptamers) and catalysis (e.g., ribozymes). Building on these foundations, researchers have begun to exploit the predictable base-pairing properties of RNA and DNA in order to utilize nucleic acids as functional materials that can undergo a molecular "switching" ...

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