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Variants of concern (VOC) of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), including alpha, beta, gamma, delta, and omicron, threaten to prolong the pandemic, leading to more global morbidity and mortality. Genome ...
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Variants of concern (VOC) of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), including alpha, beta, gamma, delta, and omicron, threaten to prolong the pandemic, leading to more global morbidity and mortality. Genome sequencing is the mainstay of tracking the evolution of the virus, but is costly, slow, and not easily accessible. Multiplex quantitative RT-PCR assays for SARS-CoV-2 have been developed that identify all VOCs as well as other mutations of interest in the viral genome, nine mutations in total, using single-nucleotide discriminating molecular beacons. The presented variant molecular beacon assays showed a limit of detection of 50 copies of viral RNA, with 100% specificity. Twenty-six SARS-CoV-2-positive patient samples were blinded and tested using a two-tube assay. When testing patient samples, the assay was in full agreement with results from deep sequencing with a sensitivity and specificity of 100% (26 of 26). We have used our design methodology to rapidly design an assay that detects the new omicron variant. This omicron assay was used to accurately identify this variant in 17 of 33 additional patient samples. These quantitative RT-PCR assays identify all currently circulating VOCs of SARS-CoV-2, as well as other important mutations in the spike protein coding sequence. These assays can be easily implemented on broadly available five-color thermal cyclers and will help track the spread of these variants.(J Mol Diagn 2022, 24: 309-319; https://doi.org/10.1016/j.jmoldx.2022.01.004)
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We have developed a simple isothermal (55 degrees C) reaction that permits detection of DNA targets using only two components: a molecular beacon and a site-specific DNA nickase without deoxyribonucleotide triphosphates and primer...
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We have developed a simple isothermal (55 degrees C) reaction that permits detection of DNA targets using only two components: a molecular beacon and a site-specific DNA nickase without deoxyribonucleotide triphosphates and primers. The loop sequence of the molecular beacon should contain a DNA nickase recognition site. The nickase-molecular beacon (NMB) combination permits a 100-fold increase in fluorescent signal. The applications of the NMB assay for enhancement of fluorescent signal in some isothermal methods are discussed. (c) 2005 Elsevier Inc. All rights reserved.
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Micro ribose nucleic acid (miRNA) recognition is of remarkable significance in the investigation of its functions as well as diagnosis of various diseases. The significance of miRNA itself is owing to the complex regulatory roles ...
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Micro ribose nucleic acid (miRNA) recognition is of remarkable significance in the investigation of its functions as well as diagnosis of various diseases. The significance of miRNA itself is owing to the complex regulatory roles in several physiological processes and its close association with important diseases such as cardiovascular problems, diabetes, Alzheimer's disease and different kinds of cancers. On the other hand, there are numerous challenges to conquer in the recognition of miRNA, consisting of low abundance, varied concentration range, small size, prolonged extraction process from cells and sequence resemblances. Traditional approaches for miRNA recognition do not meet the present demands; therefore, several fast and effective strategies have been developed with a superior focus on specificity and sensitivity such as molecular beacon (MB) probes. These probes have been widely used for miRNA monitoring with high sensitivity both in living cells and in vitro. Compared to other systems, such as qRT-PCR, studies demonstrated that MB probes can be applied as effective equipment for assaying miRNAs by taking their advantages e.g. good specificity, short hybridization period, and easy operation. Therefore, this review highlights astonishing abilities and structure of MBs and their principle. Also, the use of atomic thickness nanomaterials such as quantum dots, molybdenum disulfide, and molybdenum carbide are introduced. Moreover, new advances in developing MB approaches that are able to trace miRNA in living cells and in vivo are discussed. (C) 2020 Elsevier B.V. All rights reserved.
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Graphical abstract Display Omitted Abstract We have developed RNA-based quencher-free molecular aptamer beacons (RNA-based QF-MABs) for the detection of ATP, taking advantage of the conformational changes associated with ATP bindi...
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Graphical abstract Display Omitted Abstract We have developed RNA-based quencher-free molecular aptamer beacons (RNA-based QF-MABs) for the detection of ATP, taking advantage of the conformational changes associated with ATP binding to the ATP-binding RNA aptamer. The RNA aptamer, with its well-defined structure, was readily converted to the fluorescence sensors by incorporating a fluorophore into the loop region of the hairpin structure. These RNA-based QF-MABs exhibited fluorescence signals in the presence of ATP relative to their low background signals in the absence of ATP. The fluorescence emission intensity increased upon formation of a RNA-based QF-MAB·ATP complex.
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We have constructed a simple and efficient system-based on quencher-free molecular aptamer beacons (QF-MABs)-for probing ATP. In the absence of ATP, the fluorescence of a pyrene fluorophore on the loop position (15 nucleotides fro...
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We have constructed a simple and efficient system-based on quencher-free molecular aptamer beacons (QF-MABs)-for probing ATP. In the absence of ATP, the fluorescence of a pyrene fluorophore on the loop position (15 nucleotides from the 5' end) of the optimal QF-MAB was quenched by the neighboring nucleobases; in its presence, fluorescence was recovered, due to a conformational change in the secondary structure of the QF-MAB. (C) 2015 Elsevier Ltd. All rights reserved.
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Lung cancer is the leading cause of cancer death worldwide. There is no effective early diagnostic technology for lung cancer. microRNAs (miRNAs) are noncoding RNA molecules which regulate the process of cell growth and differenti...
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Lung cancer is the leading cause of cancer death worldwide. There is no effective early diagnostic technology for lung cancer. microRNAs (miRNAs) are noncoding RNA molecules which regulate the process of cell growth and differentiation in human cancers. Hsa-miR-155 (miR-155), highly expressed in non-small-cell lung cancer (NSCLC), can be used as a diagnostic marker for NSCLC. Dynamic observation of miR-155 is critical to diagnose NSCLC. A novel molecular beacon (MB) of miR-155 was designed to image the expression of miR-155 in NSCLC. Then miR-155 was detected in vitro by laser confocal microscopy and in vivo by stereomicroscope imaging system, respectively. The present study demonstrated that intracellular miR-155 could be successfully and quickly detected by novel miR-155 MBs. As a noninvasive monitoring approach, MBs could be used to diagnose lung cancer at early stage through molecular imaging.
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The decoder based on molecular beacon DNA computing is an indispensable hardware foundation of the DNA computer system. It has a direct influence on the development process of DNA computer technology. Since the rules of binary log...
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The decoder based on molecular beacon DNA computing is an indispensable hardware foundation of the DNA computer system. It has a direct influence on the development process of DNA computer technology. Since the rules of binary logic can also be used for switching signals in molecular converter, this paper puts forward a new implementation method of logic gates, which based on the molecular beacon calculation mode, and constructs a combinational logic circuit, 2-4 decoder. In this method, logic gates are represented by molecular beacons, signals are represented by DNA single strands, which can realize the operations of logic gates in DNA type; the connection between two logic gates according to the circuit levers can be realized by the marks of tubes. It uses including heating, elution, purification, marking and testing and some standard biological engineering technology. The construction of the molecular beacon decoder can be used for the development of DNA computer.
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A molecular beacon, an oligonucleotide probe with inherent signal transduction mechanisms, is an optimal tool for visualizing real-time mRNA hybridization in single living cells. Each molecular beacon (MB) consists of a single-str...
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A molecular beacon, an oligonucleotide probe with inherent signal transduction mechanisms, is an optimal tool for visualizing real-time mRNA hybridization in single living cells. Each molecular beacon (MB) consists of a single-stranded DNA molecule in a stem-loop conformation with a fluorophore linked to the 5' end and a quencher at the 3' end. In this study, we demonstrate realtime monitoring of mRNA-DNA hybridization inside living cells using molecular beacons. A MB specific for beta -actin mRNA has been designed and synthesized. After microinjection into the cytoplasm of single living kangaroo rat kidney cells (PtK2 cells), the MB hybridizes with beta -actin mRNA as shown by fluorescence measurements over time. Hybridization dynamics have been followed. Strict control experiments have been carried out to confirm that the fluorescence signal increase is indeed due to the hybridization of mRNA inside single living cells. Variation in the MB/mRNA hybridization fluorescent signal has been observed for different PtK2 cells, which indicates the amount of mRNA in different cells is different. We have also monitored the beta -1 andrenergic receptor mRNA inside the PtK2 cells. These studies demonstrate the feasibility of using MBs and the ultrasensitivity achieved in our fluorescence imaging system for real-time detection of mRNA hybridization and for the visualization of oligo-nucleotide/mRNA interactions inside single living cells.
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We report two novel electrochemical sensors (E-sensors) for the detection of target DNA and miRNA. The E-sensors were fabricated using label-free functional allosteric molecular beacons (aMBs), which can form streptavidin aptamers...
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We report two novel electrochemical sensors (E-sensors) for the detection of target DNA and miRNA. The E-sensors were fabricated using label-free functional allosteric molecular beacons (aMBs), which can form streptavidin aptamers to bind to streptavidin peroxidase polymer and so generate catalytic currents in the presence of the targets. These E-sensors eliminate the antigen antibody interactions which require sophisticated DNA modification. During the experiment, we found a pair of CV peaks located at around 0.17V. These peaks contributed to the redox reaction between TMB and TMB~+, and the adsorption-desorption process of TMB~+ to the negative aMB backbone. When the E-sensor was hybridized with the complement of the aMB sequence, a pair of CV peaks were found at around 0.47V which were related to the redox reaction between TMB~+ and TMB~(2+), and the process of intercalation of the planar structure of TMB~(2+) to dsDNA. The RSV-aMB E-sensor could detect 44amol RSV DNA in the 4μL sample and performed well in complicated biological environments. The let-7a-aMB E-sensor reached a detection limit of 13.6amol let-7a miRNA in the 4μL sample and showed good selectivity for one base mismatched miRNA.
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A dual-mode molecular beacon on a multiplexed substrate has been developed and applied to the measurement of unlabeled human viral RNA. The detection system is based on a combined surface-enhanced Raman scattering (SERS) and fluor...
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A dual-mode molecular beacon on a multiplexed substrate has been developed and applied to the measurement of unlabeled human viral RNA. The detection system is based on a combined surface-enhanced Raman scattering (SERS) and fluorescent molecular beacon assay that is assembled on Nanobarcodes?particles. In this assay, a molecular beacon probe terminated with a fluorescent Raman label dye is conjugated to the metallic Nanobarcodes?particles. The molecular beacon probe is a single-stranded oligonucleotide that has been designed with a hairpin structure that holds the dye at 3'-end close to the particle surface when the probe is attached through a 5'-thiol group. In this configuration, the SERS spectrum of the label was obtained and its fluorescence quenched because the dye is in very close proximity to a noble metal surface with nanoscale features (Nanobarcodes?particles). The SERS signal decreased and the fluorescence signal increased when target viral RNA was captured by this molecular beacon probe. Inaddition, a hepatitis C virus reverse transcriptase-polymerase chain reaction (HCV RT-PCR) product was detected using this dual-mode beacon. The development of a multiplexed, label-free assay system with the reassurance offered by detection of two distinctly separate signals offers significant benefits for rapid molecular diagnostics.
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