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This review is focused on antisense and functional nucleic acid used for completely rational drug design and drug target assessment, aiming to reduce the time and money spent and increase the successful rate of drug development. Nucleic acids have unique properties that play two essential roles in drug development as drug targets and as drugs. Drug targets can be messenger, ribosomal, non-coding RNAs, ribozymes, riboswitches, and other RNAs. Furthermore, various antisense and functional nucleic acids can be valuable tools in drug discovery. Many mechanisms for RNA-based control of gene expression in both pro-and-eukaryotes and engineering approaches open new avenues for drug discovery with a critical role. This review discusses the design principles, applications, and prospects of antisense and functional nucleic acids in drug delivery and design. Such nucleic acids include antisense oligonucleotides, synthetic ribozymes, and siRNAs, which can be employed for rational antibacterial drug development that can be very efficient. An important feature of antisense and functional nucleic acids is the possibility of using rational design methods for drug development. This review aims to popularize these novel approaches to benefit the drug industry and patients. [ABSTRACT FROM AUTHOR]

This review is focused on antisense and functional nucleic acid used for completely rational drug design and drug target assessment, aiming to reduce the time and money spent and increase the successful rate of drug development. Nucleic acids have unique properties that play two essential roles in drug development as drug targets and as drugs. Drug targets can be messenger, ribosomal, non-coding RNAs, ribozymes, riboswitches, and other RNAs. Furthermore, various antisense and functional nucleic acids can be valuable tools in drug discovery. Many mechanisms for RNA-based control of gene expression in both pro-and-eukaryotes and engineering approaches open new avenues for drug discovery with a critical role. This review discusses the design principles, applications, and prospects of antisense and functional nucleic acids in drug delivery and design. Such nucleic acids include antisense oligonucleotides, synthetic ribozymes, and siRNAs, which can be employed for rational antibacterial drug development that can be very efficient. An important feature of antisense and functional nucleic acids is the possibility of using rational design methods for drug development. This review aims to popularize these novel approaches to benefit the drug industry and patients.

Biomolecular engineering can be used to purposefully manipulate biomolecules, such as peptides, proteins, nucleic acids and lipids, within the framework of the relations among their structures, functions and properties, as well as their applicability to such areas as developing novel biomaterials, biosensing, bioimaging, and clinical diagnostics and therapeutics. Nanotechnology can also be used to design and tune the sizes, shapes, properties and functionality of nanomaterials. As such, there are considerable overlaps between nanotechnology and biomolecular engineering, in that both are concerned with the structure and behavior of materials on the nanometer scale or smaller. Therefore, in combination with nanotechnology, biomolecular engineering is expected to open up new fields of nanobio/bionanotechnology and to contribute to the development of novel nanobiomaterials, nanobiodevices and nanobiosystems. This review highlights recent studies using engineered biological molecules (e.g., oligonucleotides, peptides, proteins, enzymes, polysaccharides, lipids, biological cofactors and ligands) combined with functional nanomaterials in nanobio/bionanotechnology applications, including therapeutics, diagnostics, biosensing, bioanalysis and biocatalysts. Furthermore, this review focuses on five areas of recent advances in biomolecular engineering: (a) nucleic acid engineering, (b) gene engineering, (c) protein engineering, (d) chemical and enzymatic conjugation technologies, and (e) linker engineering. Precisely engineered nanobiomaterials, nanobiodevices and nanobiosystems are anticipated to emerge as next-generation platforms for bioelectronics, biosensors, biocatalysts, molecular imaging modalities, biological actuators, and biomedical applications. [ABSTRACT FROM AUTHOR]

In recent years, CRISPR-Cas (stands for: clustered regularly interspaced short palindromic repeats - CRISPR associated protein) based technologies have gained increasing attention in the biosensing field. Thanks to excellent sequence specificity, their use is of particular interest for detecting nucleic acid (NA) targets. In this context, signal generation and amplification can be realized by employing the cis -cleavage activity of the Cas9 protein, although other options involving the catalytically inactive dead Cas9 (dCas9) are increasingly explored. The latter are however mostly based on complex protein engineering processes and often lack efficient signal amplification. Here we showed for the first time that flexible signal generation and amplification properties can be integrated into the CRISPR-dCas9 complex based on a straightforward incorporation of a DNA sequence into the trans-activating CRISPR RNA (tracrRNA). The intrinsic nuclease activity of the engineered complex remained conserved, while the incorporated DNA stretch enabled two modes of amplified fluorescent signal generation: (1) as an RNA-cleaving DNA-based enzyme (DNAzyme) or (2) as hybridization site for biotinylated DNA probes, allowing subsequent enzyme labeling. Both signal generation strategies were demonstrated in solution as well as while coupled to a solid surface. Finally, in a proof of concept bioassay, we demonstrated the successful detection of single stranded DNA on magnetic microbeads using the engineered CRISPR-dCas9 complex. Thanks to the flexibility of incorporating different NA-based signal generation and amplification strategies, this novel NA engineering approach holds enormous promise for many new CRISPR-based biosensing applications. [Display omitted] • TracrRNA-DNA was engineered for integrating signal amplification with CRISPR-Cas9. • Signal amplification was enabled by coupling DNA/protein enzymes to tracrRNA-DNA. • Functionality of CRISPR complex was unaffected by the modification. • Engineered CRISPR complex generated amplified signal whilst tethered to beads. • Bead-based bioassay was developed for DNA detection using engineered CRISPR complex. [ABSTRACT FROM AUTHOR]

Nucleic acids have proven to be a very suitable medium for engineering various nanostructures and devices. While synthetic DNAs are commonly used for self-assembly of nanostructures and devices in vitro, functional RNAs, such as ribozymes, are employed both in vitro and in vivo. Allosteric ribozymes have applications in molecular computing, biosensoring, high-throughput screening arrays, exogenous control of gene expression, and others. They switch on and off their catalytic function as a result of a conformational change induced by ligand binding. Designer ribozymes are engineered to respond to different effectors by in vitro selection, rational and computational design methods. Here, I present diverse computational methods for designing allosteric ribozymes with various logic functions that sense oligonucleotides or small molecules. These methods yield the desired ribozyme sequences within minutes in contrast to the in vitro selection methods, which require weeks. Methods for synthesis and biochemical testing of ribozymes are also discussed. [ABSTRACT FROM AUTHOR]

Summary: Freestanding ultrathin nano-membranes (FUN-membranes) are two-dimensional materials of nanoscale thickness (less than 100nm) with little or no substrate support. Recently, there has been an emerging interest to rationally design such membranes for a broad spectrum of appealing applications ranging from lightweight, foldable electronic devices to highly efficient separation and sensing systems. In this review, we aim to summarize the recent developments of FUN-membranes fabricated via an assortment of bottom-up self-assembly strategies. In particular, we will describe the various fabrication methods, characterization approaches, as well as the properties and applications of FUN-membrane materials. [Copyright &y& Elsevier]

• Material science at the nanoscale provides new standards in pathogen diagnostics. • New materials may overcome the limitations of dominant COVID-19 diagnostics. • Distinct classes of nanomaterials and nanochemistry enable new diagnostic solutions. The COVID-19 pandemic revealed fundamental limitations in the current model for infectious disease diagnosis and serology, based upon complex assay workflows, laboratory-based instrumentation, and expensive materials for managing samples and reagents. The lengthy time delays required to obtain test results, the high cost of gold-standard PCR tests, and poor sensitivity of rapid point-of-care tests contributed directly to society's inability to efficiently identify COVID-19-positive individuals for quarantine, which in turn continues to impact return to normal activities throughout the economy. Over the past year, enormous resources have been invested to develop more effective rapid tests and laboratory tests with greater throughput, yet the vast majority of engineering and chemistry approaches are merely incremental improvements to existing methods for nucleic acid amplification, lateral flow test strips, and enzymatic amplification assays for protein-based biomarkers. Meanwhile, widespread commercial availability of new test kits continues to be hampered by the cost and time required to develop single-use disposable microfluidic plastic cartridges manufactured by injection molding. Through development of novel technologies for sensitive, selective, rapid, and robust viral detection and more efficient approaches for scalable manufacturing of microfluidic devices, we can be much better prepared for future management of infectious pathogen outbreaks. Here, we describe how photonic metamaterials, graphene nanomaterials, designer DNA nanostructures, and polymers amenable to scalable additive manufacturing are being applied towards overcoming the fundamental limitations of currently dominant COVID-19 diagnostic approaches. In this paper, we review how several distinct classes of nanomaterials and nanochemistry enable simple assay workflows, high sensitivity, inexpensive instrumentation, point-of-care sample-to-answer virus diagnosis, and rapidly scaled manufacturing. [ABSTRACT FROM AUTHOR]

We demonstrate the first biosensing strategy that relies on quantum dot (QD) fluorescence blinking to report the presence of a target molecule. Unlike other biosensors that utilize QDs, our method does not require the analyte to induce any fluorescence intensity or color changes, making it readily applicable to a wide range of target species. Instead, our approach relies on the understanding that blinking, a single particle phenomenon, is obscured when several QDs lie within the detection volume of a confocal microscope. If QDs are engineered to aggregate when they encounter a particular target molecule, the observation of quasi-continuous emission should indicate its presence. As proof of concept, we programmed DNAs to drive rapid isothermal assembly of QDs in the presence of a target strand (oncogene K- ras ). The assemblies, confirmed by various gel techniques, contained multiple QDs and were readily distinguished from free QDs by the absence of blinking., Competing Interests: Notes The authors declare no competing financial interest.

We describe design parameters for the synthesis and analytical application of a label-free RNA molecular beacon, termed Spinach.ST. The RNA aptamer Spinach fluoresces upon binding the small-molecule fluorophore DFHBI ((Z)-4-(3,5-difluoro-4-hydroxybenzylidene)-1,2-dimethyl-1H-imidazol-5(4H)-one). Spinach has been reengineered by extending its 5'- and 3'-ends to create Spinach.ST, which is predicted to fold into an inactive conformation that fails to bind DHFBI. Hybridization of a trigger oligonucleotide to a designed toehold on Spinach.ST initiates toehold-mediated strand displacement and restores the DFHBI-binding, fluorescence-enhancing conformation of Spinach. The versatile Spinach.ST sensor can detect DNA or RNA trigger sequences and can readily distinguish single-nucleotide mismatches in the trigger toehold. Primer design techniques are described that augment amplicons produced by enzymatic amplification with Spinach.ST triggers. Interaction between these triggers and Spinach.ST molecular beacons leads to the real-time, sequence-specific quantitation of these amplicons. The use of Spinach.ST with isothermal amplification reactions such as nucleic acid sequence-based amplification (NASBA) may enable point-of-care applications. The same design principles could also be used to adapt Spinach reporters to the assay of nonnucleic acid analytes in trans., (© 2015 Elsevier Inc. All rights reserved.)

We have re-engineered the fluorescent RNA aptamer Spinach to be activated in a sequence-dependent manner. The original Spinach aptamer was extended at its 5'- and 3'-ends to create Spinach.ST, which is predicted to fold into an inactive conformation and thus prevent association with the small molecule fluorophore DFHBI. Hybridization of a specific trigger oligonucleotide to a designed toehold leads to toehold-initiated strand displacement and refolds Spinach into the active, fluorophore-binding conformation. Spinach.ST not only specifically detects its target oligonucleotide but can discriminate readily against single-nucleotide mismatches. RNA amplicons produced during nucleic acid sequence-based amplification (NASBA) of DNA or RNA targets could be specifically detected and reported in real-time by conformational activation of Spinach.ST generated by in vitro transcription. In order to adapt any target sequence to detection by a Spinach reporter we used a primer design technique that brings together otherwise distal toehold sequences via hairpin formation. The same techniques could potentially be used to adapt common Spinach reporters to non-nucleic acid analytes, rather than by making fusions between aptamers and Spinach., (© 2014 Bhadra and Ellington; Published by Cold Spring Harbor Laboratory Press for the RNA Society.)