Importantly, the hairpin structure of the probe is responsible for both the low fluorescent background and improved selectivity. Furthermore, the signal is generated in a reversible manner; thus, if the analyte is removed, the signal is reduced to the background. This paper highlights the advantages of MB probes and discusses the approaches that address the challenges in MB probe design. A review: microRNA detection methods.
The importance of miRNA itself is due to the complicated regulatory functions it plays in various life processes and its close relation with some diseases. Traditional methods for miRNA detection do not meet the current demands, so various novel methods have been developed with a special focus on sensitivity and specificity. Herein, the authors summarize and discuss the newly developed miRNA detection methods.
We describe here a new method for highly efficient detection of microRNAs by northern blot anal. By modifying DNA oligonucleotides with LNAs using a design, in which every third nucleotide position was substituted by LNA, the authors could use the probes in northern blot anal. The sensitivity in detecting mature microRNAs by northern blots was increased by at least fold compared to DNA probes, while simultaneously being highly specific, as demonstrated by the use of different single and double mismatched LNA probes. This review highlights various methods that can be used for a sensitive detection of nucleic acids without using thermal cycling procedures, as is done in PCR or LCR.
Isothermal Amplification of Nucleic Acids. Isothermal amplification of nucleic acids is a simple process that rapidly and efficiently accumulates nucleic acid sequences at const. Since the early s, various isothermal amplification techniques have been developed as alternatives to polymerase chain reaction PCR. These isothermal amplification methods have been used for biosensing targets such as DNA, RNA, cells, proteins, small mols.
The applications of these techniques for in situ or intracellular bioimaging and sequencing have been amply demonstrated. Amplicons produced by isothermal amplification methods have also been utilized to construct versatile nucleic acid nanomaterials for promising applications in biomedicine, bioimaging, and biosensing. The integration of isothermal amplification into microsystems or portable devices improves nucleic acid-based on-site assays and confers high sensitivity.
Single-cell and single-mol. In this review, we provide a comprehensive overview of the isothermal amplification of nucleic acids encompassing work published in the past two decades. First, different isothermal amplification techniques are classified into three types based on reaction kinetics. Then, we summarize the applications of isothermal amplification in bioanal.
Finally, several challenges and perspectives in the field are discussed. Roll with it: The quant. A novel method for miRNA anal. Real-time detection of isothermal amplification reactions with thermostable catalytic hairpin assembly. Catalytic hairpin assembly CHA is an enzyme-free amplification method that has previously proven useful in amplifying and transducing signals at the terminus of nucleic acid amplification reactions.
CHA circuits were designed and optimized for both high- and low-temp. The resulting circuits not only increased the specificity of detection but also improved the sensitivity by as much as to fold over comparable real-time detection methods. These methods have been condensed into a set of general rules for the design of thermostable CHA circuits with high signals and low noise.
A simple electrochemical biosensor for highly sensitive and specific detection of microRNA based on mismatched catalytic hairpin assembly. MicroRNAs miRNAs play vital regulatory roles in cancer development and a variety of diseases, which make them become promising biomarkers. Here, a simple electrochem.
Under the optimal exptl. It was also applied to the detn. Thus, this biosensing strategy might become a potential alternative tool for detection of miRNA in biomedical research and early clin. Hybridization chain reaction amplification of microRNA detection with a tetrahedral DNA nanostructure-based electrochemical biosensor. There remains a great challenge in the sensitive detection of microRNA because of the short length and low abundance of microRNAs in cells.
Here, the authors demonstrated an ultrasensitive detection platform for microRNA by combining the tetrahedral DNA nanostructure probes and hybridization chain reaction HCR amplification. Compared to the widely used supersandwich amplification, the detection limits are improved by 3 orders of magnitude. The uncontrolled surface immobilization and consumption of target mols. Taking advantage of DNA nanotechnol.
In situ DNA-templated synthesis of silver nanoclusters for ultrasensitive and label-free electrochemical detection of microRNA. ACS Appl. Interfaces , 7 , — , DOI: On the basis of the use of silver nanoclusters AgNCs in situ synthesized by cytosine C -rich loop DNA templates as signal amplification labels, the development of a label-free and highly sensitive method for electrochem.
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These surface-captured intermediate sequences further trigger the hybridization chain reaction HCR amplification to form dsDNA polymers with numerous C-rich loop DNA templates on the electrode surface. Featured with high sensitivity and label-free capability, the proposed sensing scheme can thus offer new opportunities for achieving sensitive, selective, and simple detection of different types of microRNA targets.
A plasmonic colorimetric strategy for visual miRNA detection based on hybridization chain reaction. In this work, a novel colorimetric strategy for miRNA anal. The stability variation of the colloid system can then be monitored by recording corresponding UV-vis spectrum and initial miRNA level is thus detd. This sensing system involves only four DNA strands which is quite simple. The practical utility is confirmed to be excellent by employing different biol. Ed , 56 , — , DOI: We rationally engineered an elegant entropy-driven DNA nanomachine with three-dimensional track and applied it for intracellular miRNAs imaging.
The proposed nanomachine is activated by target miRNA binding to drive a walking leg tethered to gold nanoparticle with a high d.
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The autonomous and progressive walk on the DNA track via the entropy-driven catalytic reaction of intramol. Our nanomachine outperforms the conventional intermol. Moreover, the nanomachine was applied for miRNA imaging inside living cells. ACS Sens. Given the low miRNA expression at the earlier stage of diseases, its amplified detection still requires more efforts. Inspired by the two-stage arithmetic amplifier of elec. Here the catalytically inactive DNAzyme subunits were resp. The present CHA-DNAzyme amplifier can be employed as a versatile and general sensing platform for analyzing other analytes e.
Given the attractive anal. Increasing prevalence of pollen allergies has raised concerns about human health. Development of a facile and precise method to detect pollen allergens would thus be of significance for environmental assessments and medical diagnoses. Here we report a sensitive colorimetric method to detect the Japanese cedar pollen allergen, Cry j 2. The method consists of two steps: a signal amplification based on the catalytic DNA hairpin self-assembly, followed by a signal transduction using the salt-induced non-crosslinking aggregation of gold nanoparticles densely modified with short DNA.
The assay exhibits a detection limit of 0. Moreover, the assay enables the detection of Cry j 2 spiked in soil solns. The signal amplification system includes an anti-Cry j 2 DNA aptamer, which accounts for the absence of false responses to five nontarget allergen proteins. The present method could be of general applicability to various proteins by using appropriate aptamers.
Highly sensitive and specific electrochemical biosensor for microRNA detection by coupling catalytic hairpin assembly with rolling circle amplification. Analyst , , — , DOI: Background: MicroRNA plays a significant role in gene regulation and is usually regarded as an important biol. Methods: In this expt. Results: This strategy has a good linear range of 0.
Conclusion: This method can be applied not only for microRNA detection with high sensitivity and speed, but can also detect small mols.
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Sensitive detection of miRNA by using hybridization chain reaction coupled with positively charged gold nanoparticles. Under optimal conditions, miRNA detection could be realized in the range of 20 pM nM with a detection limit of 6. Importantly, the assay could realize the detection of miRNA in human serum samples. An electrochemical biosensor for sensitive detection of microRNA combining target recycling with cascade catalysis for signal amplification. In this work, a new electrochem.
It is worth pointing out that target recycling was achieved only based on strand displacement process without the help of nuclease. Moreover, porous TiO2 nanosphere was synthesized, which could offer more surface area for Pt nanoparticles PtNPs enwrapping and enhance the amt. With the mimicking sandwich-type reaction, the cascade catalysis amplification strategy was carried out by AOx catalyzing ethanol to acetaldehyde with the concomitant formation of high concn.
This newly designed biosensor provided a sensitive detection of miRNA from 0. Spandidos Publications Ltd. Sanger mibase, miRanda and TargetScan were used to identify common predicted genes. Databases were used to identify homologous sequences on mRNAs of putative target genes that may be directly bound by the miRNAs identified. We found that cytokines and epigenetic regulators may be putative target genes of these miRNAs. Using ingenuity pathway anal. Molecular beacon-based bioimaging of multiple microRNAs during myogenesis. Biomaterials , 32 , — , DOI: Elsevier Ltd. C2C12 cells were transfected with an MB targeting miRa and contg.
In vitro and in vivo fluorescence anal. Influenza A virus isolation, culture and identification. Influenza A viruses IAVs cause epidemics and pandemics that result in considerable financial burden and loss of human life. To manage annual IAV epidemics and prep. Fundamental techniques essential for procuring such knowledge are IAV isolation and culture from exptl.
Topic Collection: Influenza Epidemic/ Pandemic
Here the authors present a detailed protocol for IAV sample collection and processing, amplification in chicken eggs or mammalian cells, and identification from samples contg. This protocol is robust, and it allows for the generation of virus cultures that can be used for downstream analyses. Once exptl. Increased time frames may be required for less experienced lab. BMC Microbiol. BioMed Central Ltd.
Background: Avian influenza remains a serious threat to human health. The consequence of human infection varies markedly among different subtypes of avian influenza viruses. This study aims at elucidating how avian influenza infection perturbs the host's miRNA regulatory pathways that may lead to adverse pathol. Results: The results showed that dysregulation of miRNA expression was mainly obsd. We found that miR, miR, miR, miRp, miRa and miRb were differentially expressed in response to influenza A virus infection.
This was supported by the observation that the inhibitory effect could be reversed by antagomiR Quantitative polymerase chain reaction qPCR was used to validate the profiling results in the discovery set and in a validation set of 31 controls and 22 patients with early-stage NSCLC. The utility of miR and miRp serum-based biomarkers as minimally invasive screening and triage tools for subsequent diagnostic evaluation warrants additional validation. PLoS Genet. We identified miR as a neg. Consistently with the in vitro studies, mouse xenograft studies validated that miR suppressed NSCLC tumor growth in vivo.
Pandemic Influenza (Modular Texts Series)
Overall, these findings identify the dual inhibition of HO-1 by miR as a novel functional mechanism of miRNA, which results in a more effective inhibition of oncogenic mRNA, and leads to a tumor suppressive effect. MicroRNA regulation of human protease genes essential for influenza virus replication.
Meliopoulos, Victoria A. Keegan; Tompkins, S. Mark; Tripp, Ralph A. Influenza A virus causes seasonal epidemics and periodic pandemics threatening the health of millions of people each year. Vaccination is an effective strategy for reducing morbidity and mortality, and in the absence of drug resistance, the efficacy of chemoprophylaxis is comparable to that of vaccines. However, the rapid emergence of drug resistance has emphasized the need for new drug targets.
Knowledge of the host cell components required for influenza replication has been an area targeted for disease intervention.
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In this study, the human protease genes required for influenza virus replication were detd. The genes validated as crit. Analyses of host microRNAs predicted to govern expression of these genes showed that eight miRNAs regulated gene expression during virus replication. These findings identify unique host genes and microRNAs important for influenza replication providing potential new targets for disease intervention strategies.
Insights , 8 , — , DOI: Kumar, Amod; Asaf, V. Libertas Academica Ltd. Highly pathogenic Avian influenza HPAI is a notifiable viral disease caused by avian influenza type A viruses of the Orthomyxoviridae family. Type A influenza genome consists of eight segments of neg. Since these proteins are crit. PLoS Pathog. Hendra and Nipah viruses family Paramyxoviridae, genus Henipavirus are bat-borne viruses that cause fatal disease in humans and a range of other mammalian species.
Gaining a deeper understanding of host pathways exploited by henipaviruses for infection may identify targets for new anti-viral therapies. Here we have performed genome-wide highthroughput agonist and antagonist screens at biosafety level 4 to identify host-encoded microRNAs miRNAs impacting henipavirus infection in human cells. MiR also promoted Nipah virus infection, but did not affect infection by paramyxoviruses from other genera, indicating specificity in the virus-host interaction.
Infection promotion was primarily mediated via the ability of miR to significantly enhance henipavirus-induced membrane fusion. Cell signalling receptors of ephrins, namely EphA5 and EphA7, were identified as novel neg. The expression of these receptors, as well as EphB4, were suppressed by miR overexpression, suggesting that simultaneous inhibition of several Ephs by the miRNA contributes to enhanced infection and fusion. Immune-responsive miR levels was also up-regulated in the biofluids of ferrets and horses infected with Hendra virus, suggesting that the host innate immune response may promote henipavirus spread and exacerbate disease severity.
This study is the first genome-wide screen of miRNAs influencing infection by a clin. Viruses, microRNAs, and host interactions. Annual Reviews Inc. These regulatory RNAs provide a unique level of posttranscriptional gene regulation that modulates a range of fundamental cellular processes. Several viruses, esp. Current evidence indicates that viruses use these miRNAs to manipulate both cellular and viral gene expression.
Here we discuss our current knowledge of viral miRNAs and virally influenced cellular miRNAs and their relationship to viral infection. A targeted, self-delivered, and photocontrolled molecular beacon for mRNA detection in living cells.
The spatiotemporal dynamics of specific mRNA mols. To solve this problem, we have developed a targeted, self-delivered, and photocontrolled aptamer-based mol. A light activation strategy was employed by inserting two photolabile groups in the CP sequence, enabling control over the MB's intracellular function. After the probe was guided to the target cell via specific binding of aptamer AS to nucleolin on the cell membrane, light illumination released the MB for mRNA monitoring.
Consequently, the MB is able to perform live-cell mRNA imaging with precise spatiotemporal control, while the CP acts as both a tracer for intracellular distribution of the MB before photoinitiation and an internal ref. Rationally designed molecular beacons for bioanalytical and biomedical applications. Nucleic acids hold promise as biomols. Their well-defined structures and compns. Beyond the Bump A clinical psychologist's guide to navigating t Pretty Unhealthy.
Inflamed Mind A radical new approach to depression. Where Did My Libido Go? It's About Bloody Time. Can You Die of a Broken Heart? A heart surgeon's insight into what makes us tick. Healthy Hormones A practical guide to balancing your hormones. The Nude Nutritionist. The Complete Guide to Asperger's Syndrome. The Postnatal Depletion Cure A complete guide to rebuilding your health and Curr Neurol Neurosci Rep ; The successful use of extra-corporeal membrane oxygenation in the management of a pregnant woman with severe H1N1 influenza complicated by pneumonitis and adult respiratory distress syndrome.
Int J Obstet Anesth ; Crit Care ; H1N1 infection and the kidney in critically ill patients. J Nephrol ; H1N1 influenza A virus-associated acute lung injury: response to combination oseltamivir and prolonged corticosteroid treatment. Intensive Care Med ; Ministry of Health, Government of India. Clinico-epidemiological features of the hospitalized patients with pandemic influenza A H1N1 virus infection in Saurashtra region, India September, to February, An insight into the swine-influenza A H1N1 virus infection in human.
Non-invasive ventilation in acute respiratory failure due to H1N1 influenza. This article has been cited by. Devanshi J. Access Statistics.