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A novel method of catalytic hairpin assembly (CHA) induced dual signal enhancement is developed for rapid detection of miRNA based on fluorescence light-up silver nanocluster (Ag NC). By the hybridization of a hairpin DNA and a single-stranded DNA, a unique probe is firstly designed. In the terminals of this probe, DNA-Ag NCs can be formed and display very weak fluorescence. In the presence of the target miRNA, the reaction of CHA can be triggered, forming two kinds of double-stranded complexes, in which the terminal DNA-Ag NCs are in close proximity to G-rich overhangs and the fluorescent signal can be dramatically enhanced. Compared with many other enzyme-based amplification strategies, this one exhibits distinct advantages of simplicity in experimental operation and a rapid detection process (within 1 h). Moreover, this assay exhibits an excellent selectivity and is successfully applied in the detection of miRNAs in complex biological media, which confirms the reliability and practicality of this protocol., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Exosomes are membrane-enclosed phospholipid extracellular vesicles. In spite of their great promise as noninvasive biomarkers for cancer diagnosis, sensitive detection of exosomes is still challenging. Herein, the detection of exosomes was changed to the detection of DNA after recognition of exosomes with its aptamers. CD63 aptamer and EpCAM aptamer were used for the detection of MCF-7 cell-secreted exosome. The recognition process was amplified through the movements of a three-dimensional DNA walker. And then, Exonuclease III- assisted electrochemical ratiometric sensor was applied for further signal amplification. Under optimal conditions, the detection limit of 1.3 × 10 4 particles/mL was obtained with excellent selectivity. Furthermore, clinical application test for the detection of exosomes in human serum was also verified.

A photocathode is described for the determination of microRNA-21 by using CuInS 2 as an active photocathode material. Exonuclease III assisted target recycling amplification was employed to enhance the detection sensitivity. The TATA-binding protein (TBP) was applied to enhance steric hindrance which decreases the photoelectrochemical intensity. This strategy is designed by combining the anti-interference photocathode material, enzyme assisted target recycling amplification and TBP induced signal off, showing remarkable amplification efficiency. Under the optimized conditions, the detection limit for microRNA-21 is as low as 0.47 fM, and a linear range was got from 1.0 × 10 -15  M to 1.0 × 10 -6  M. Graphical abstract Schematic representation of sensitive photoelectrochemical detection of microRNA-21.CuInS 2 is used as an active photocathode material. Combined Exonuclease III assisted target recycling amplification and TATA-binding protein decreased of photoelectrochemical intensity, the detection limit was 0.47 fM with good selectivity. (miR-21: microRNA-21; CS: chitosan).

The Raman probe plays an essential role in sensitive surface-enhanced Raman scattering (SERS) assay. Here, a novel Raman probe was developed by assembling gold nanoparticles in triangular pyramid DNA (TP-Au NPs). Such probe with intense electromagnetic hot spots can provide dramatically enhanced Raman scattering. Through assembling recognition DNA on one corner of the TP-DNA, the recognition event is definite and designable. The probe was characterized through TEM, and its SERS superiority was investigated. As models, circulating tumor cells and exosomes were detected with high sensitivity and selectivity by using this probe. Meanwhile, the developed SERS probe can also perform well in real world samples.

An electrochemical method is described for the sensitive detection of the activity of the enzyme T4 polynucleotide kinase (PNK) by using a DNA functionalized porphyrinic metal-organic framework (L/(Fe-P)n-MOF). In the presence of PNK, the hairpin oligonucleotide (HP1) becomes phosphorylated, and the trigger is released by lambda exonuclease (λ exo). The trigger DNA hybridizes with hairpin probe (immobilized on the gold electrode) to form a nicking endonuclease cleavage site. Thus, a single-strand capture probe is employed to hybridize with L/(Fe-P)n-MOF. The (Fe-P)n-MOF is a peroxidase mimicking material with high catalytic efficiency. By using this amplification strategy, an electrochemical signal is procured that allows for the determination of T4 PNK in the 1.0 mU·mL -1 to 1.0 U·mL -1 with a detection limit of 0.62 mU·mL -1 . The method is selective and can be used to screen for enzyme inhibitors. Conceivably, the (Fe-P)n-MOF can also be used to detect other analytes via its peroxidase-mimicking activity. Graphical abstract Schematic presentation of T4 polynucleotide kinase (PNK) detection. Two hairpin DNAs (HP) and a porphyrinic metal-organic framework with peroxidase-mimicking activity are used. The detection limit is 0.62 mU mL -1 with enzyme assisted signal amplification. This method is selective and can be used to screen for enzyme inhibitors.

Detection of ultralow concentrations of nucleic acid sequences is a central challenge in the early diagnosis of genetic diseases. Herein, we developed a target-triggering cascade multiple cycle amplification for ultrasensitive DNA detection using quartz crystal microbalance (QCM) and surface plasmon resonance (SPR). It was based on the exonuclease Ⅲ (Exo Ⅲ)-assisted signal amplification and the hybridization chain reaction (HCR). The streptavidin-coated Au-NPs (Au-NPs-SA) were assembled on the HCR products as recognition element. Upon sensing of target DNA, the duplex DNA probe triggered the Exo Ⅲ cleavage process, accompanied by generating a new secondary target DNA and releasing target DNA. The released target DNA and the secondary target DNA were recycled. Simultaneously, numerous single strands were liberated and acted as the trigger of HCR to generate further signal amplification, resulting in the immobilization of abundant Au-NPs-SA on the gold substrate. The QCM sensor results were found to be comparable to that achieved using a SPR sensor platform. This method exhibited a high sensitivity toward target DNA with a detection limit of 0.70 fM. The high sensitivity and specificity make this method a great potential for detecting DNA with trace amounts in bioanalysis and clinical biomedicine., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Cation-exchange-based chemiluminescence amplification (CXCLAmp) was developed in this study. After amplification by rolling circle amplification (RCA), the proposed RCA-CXCLAmp strategy was used to detect miRNA sensitively.

We report a strategy for the transduction of DNA hybridization into a readily detectable photoelectrochemical signal by means of a conformational change analogous to electrochemical DNA (E-DNA) approach. To demonstrate the effect of distance change for photosensitizer to the surface of electrode on the change of photocurrent, photosensitizer Ru(bpy)(2)(dcbpy)(2+) tagged DNA stem-loop structures were self-assembled onto a nanogold modified ITO electrode. Hybridization induced a large conformational change in DNA structure, which in turn significantly altered the electron-transfer tunneling distance between the electrode and photosensitizer. The resulting change in photocurrent was proportional to the concentration of DNA in the range of 1.0×10(-10)-8.0×10(-9)M. In order to improve the sensitivity of the photoelectrochemical biosensor, an amplified detection method based on isothermal strand displacement polymerization reaction was employed. With multiple rounds of isothermal strand replication, which led to strand displacement and constituted consecutive signal amplification, a detection limit of 9.4×10(-14)M target DNA was achieved., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Physical light source is absolutely necessary for usual photoelectrochemical measurement. In this work, chemiluminescence reaction rather than physical light source was used for the development of a novel photoelectrochemical DNA biosensor. CIPO (bis(2,4,5-trichlro-6-n-pentoxycarbonylphenyl)oxalate)-H(2)O(2)-9,10-diphenylanthrancene was selected as a CL system, which can produce appropriate exciting light and excite photoelectro active materials Ru(bpy)(2)dppz(2+) intercalated into the double-stranded DNA. Using such simple intercalation method, a detection limit of 4.5×10(-9) M target DNA was achieved without any amplification process. In addition, the selected CL system could be used to excite AuNPs-Ru(bpy)(2)dppz(2+) complex as well as CdSe QD multilayer, which indicated a good applicability for the established method., (Copyright © 2010 Elsevier B.V. All rights reserved.)

A novel "signal-on" assay for sensitive and selective detection of Hg(2+) in aqueous solution based on mismatched DNA and a molecular light switch complex, [Ru(phen)(2)(dppz)](2+), was reported in the present work. This Hg(2+) specific sensor system composed of two label-free DNA probes with eight T-T mismatches, which could form stable DNA duplexes by thymine-Hg(2+)-thymine (T-Hg(2+)-T) in the presence of Hg(2+). The luminescence of [Ru(phen)(2)(dppz)](2+) is very weak in aqueous solution, and significant luminescence can be observed when intercalating into DNA duplexes. By monitoring Hg(2+)-dependent luminescence intensity enhancement, highly sensitive and selective determination of Hg(2+) has been achieved. Under the optimum conditions, the luminescence intensity was proportional to the concentration of Hg(2+) in the range of 1.0x10(-9)-1.5x10(-7) M. A detection limit of 3.5x10(-10) M Hg(2+) was achieved using 3sigma. The presence of other metal ions did not interfere with the detection of Hg(2+) even at high concentrations. This approach is simple and rapid, which can be used to monitor the Hg(2+) concentration in drinking water and natural media within 30 min., (Copyright 2009 Elsevier B.V. All rights reserved.)

Both nickel and cadmium compounds have been established as group I carcinogens for several decades. Despite over-whelming evidence of these compounds’ carcinogenicity in humans, the specific underlying molecular mechanisms that govern metal induced cellular transformation remain unclear. In this study, we found that there were slightly different effects on decreased SLBP mRNA and protein as well as increased polyA H3.1 in our nickel exposed cells. This suggested that nickel and arsenic have similar effects on canonical histone mRNA transcription and translation. We also saw that the depletion of SLBP protein was reversed by inhibiting the proteosome. Finally, we showed that inhibiting the SLBP mRNA and protein levels were rescued by epigenetic modifiers suggesting that nickel’s effects on SLBP may be mediated via epigenetic mechanisms. Taken together these results suggest a similar mechanism by which both arsenic and nickel may exert their carcinogenic effects. [ABSTRACT FROM AUTHOR]

The environmental and occupational carcinogen Hexavalent Chromium (Cr(VI)) has been shown to cause lung cancer in humans when inhaled. In spite of a considerable research effort, the mechanisms of Cr(VI)-induced carcinogenesis remain largely unknown. Nupr1 (nuclear protein 1) is a small, highly basic, and unfolded protein with molecular weight of 8,800 daltons and is induced by a variety of stressors. Studies in animal models have suggested that Nupr1 is a key factor in the development of lung and pancreatic cancers, with little known about the underlying molecular mechanisms. Here we report that the level of Nupr1 is significantly increased in human bronchial epithelial BEAS2B cells following exposure to Cr(VI) through epigenetic mechanisms. Interestingly, Cr(VI) exposure also results in the loss of acetylation at histone H4K16, which is considered a ‘hallmark’ of human cancer. Cr(VI)-induced reduction of H4K16 acetylation appears to be caused by the induction of Nupr1, since (a) overexpression of Nupr1 decreased the levels of both H4K16 acetylation and the histone acetyltransferase MOF (male absent on the first; also known as Kat8, Myst 1), which specifically acetylates H4K16; (b) the loss of acetylation of H4K16 upon Cr(VI) exposure is greatly compromised by knockdown of Nupr1. Moreover, Nupr1-induced reduction of H4K16 acetylation correlates with the transcriptional down-regulation at several genomic loci. Notably, overexpression of Nupr1 induces anchorage-independent cell growth and knockdown of Nupr1 expression prevents Cr(VI)-induced cell transformation. We propose that Cr(VI) induces Nupr1 and rapidly perturbs gene expression by downregulating H4K16 acetylation, thereby contributing to Cr(VI)-induced carcinogenesis. [ABSTRACT FROM AUTHOR]

Ultraviolet radiation (UVR) from sunlight is the primary effector of skin DNA damage. Chromatin remodeling and histone post-translational modification (PTM) are critical factors in repairing DNA damage and maintaining genomic integrity, however, the dynamic changes of histone marks in response to solar UVR are not well characterized. Here we report global changes in histone PTMs induced by solar simulated UVR (ssUVR). A decrease in lysine acetylation of histones H3 and H4, particularly at positions of H3 lysine 9, lysine 56, H4 lysine 5, and lysine 16, was found in human keratinocytes exposed to ssUVR. These acetylation changes were highly associated with ssUVR in a dose-dependent and time-specific manner. Interestingly, H4K16ac, a mark that is crucial for higher order chromatin structure, exhibited a persistent reduction by ssUVR that was transmitted through multiple cell divisions. In addition, the enzymatic activities of histone acetyltransferases were significantly reduced in irradiated cells, which may account for decreased global acetylation. Moreover, depletion of histone deacetylase SIRT1 in keratinocytes rescued ssUVR-induced H4K16 hypoacetylation. These results indicate that ssUVR affects both HDAC and HAT activities, leading to reduced histone acetylation. [ABSTRACT FROM AUTHOR]

Abstract: A novel “signal-on” assay for sensitive and selective detection of Hg2+ in aqueous solution based on mismatched DNA and a molecular light switch complex, [Ru(phen)2(dppz)]2+, was reported in the present work. This Hg2+ specific sensor system composed of two label-free DNA probes with eight T–T mismatches, which could form stable DNA duplexes by thymine–Hg2+–thymine (T–Hg2+–T) in the presence of Hg2+. The luminescence of [Ru(phen)2(dppz)]2+ is very weak in aqueous solution, and significant luminescence can be observed when intercalating into DNA duplexes. By monitoring Hg2+-dependent luminescence intensity enhancement, highly sensitive and selective determination of Hg2+ has been achieved. Under the optimum conditions, the luminescence intensity was proportional to the concentration of Hg2+ in the range of 1.0×10−9–1.5×10−7 M. A detection limit of 3.5×10−10 M Hg2+ was achieved using 3σ. The presence of other metal ions did not interfere with the detection of Hg2+ even at high concentrations. This approach is simple and rapid, which can be used to monitor the Hg2+ concentration in drinking water and natural media within 30min. [Copyright &y& Elsevier]

Abstract: A sensitive label-free bio-barcode assay provided a PCR-free method for quantitative detection of two nucleic acid targets (HTLV-I and HTLV-II) simultaneously. This DNA biosensor was fabricated with two-component oligonucleotide-modified gold nanoparticles (AuNPs) and two-component oligonucleotide-modified magnetic beads (MBs), which can sandwich a specific target. After liberating the adsorbed thiolated barcode DNA strands (poly A and poly G) from the AuNPs surface with dithiothreitol (DTT) and acidic dipurinization, the electrochemical measurements were directly performed based on the redox activity of guanine (G) and adenine (A) nucleobases. Under the optimal assembling and detection conditions, a good linearity for simultaneous detection was obtained in the range from 4.4×10−11 to 2.0×10−9 M, and the detection limit (3σ) was estimated to be 1.71×10−12 M for T1-DNA and 1.55×10−12 M for T2-DNA. [Copyright &y& Elsevier]

This work develops a novel photoelectrochemical sensor for the detection of carcinoembryonic antigen (CEA) based on the composite of UCNPs with semiconductors and conformational changes in the DNA structure. Firstly, SnS 2 , ZnIn 2 S 4 and UCNPs were assembled on the surface of the ITO electrode. Then Au NPs were dropped, which could facilitate the coupling of CdSe NPs modified DNA1 via Au–S bond, giving an ITO/SnS 2 /ZnIn 2 S 4 /UCNPs/CdSe heterojunction structure. When irradiated with 980 nm near-infrared (NIR) light, the UV–visible light emitted by the UCNPs could excite the nanocomposite, producing an enhanced photoelectric reaction. Subsequently, CEA aptamer and DNA2-modified SiO 2 were added to form a Y-shaped DNA structure. At this time, the photocurrent was significantly reduced by the combination of the light-blocking effect of SiO 2 and the departure of CdSe NPs from the electrode surface. When the target CEA was added, the recognition between CEA and the aptamer led to the collapse of the Y-shaped DNA structure, the restoration of hairpin DNA and the proximity of CdSe to the electrode. Accordingly, the photocurrent signals enhanced again. Under optimal experimental conditions, the detection limit as low as 0.3 pg mL−1 was obtained with good selectivity, achieving a sensitive "on-off-on" photoelectrochemical sensor for CEA detection. [ABSTRACT FROM AUTHOR]

The analysis of DNA methylation and MTase activities is very important in the early clinical diagnosis of cancer, on purposes of providing insights into the mechanism of gene repression and developing novel drugs of treating methylation-related diseases. Combining the dendritic rolling circle amplification and Mg 2+ -dependent DNAzyme with a function of catalyzing the generation of a fluorophore-labeled nucleic acid acting as readout signal for the analyses, a new fluorescent method for DNA methyltransferase detection was reported. In the presence of DNA methyltransferases (MTase), the methylation-responsive sequence of double-stranded DNA probe was methylated and then cleaved by the methylation-sensitive restriction endonuclease Dpn I. The cleaved hybrid DNA probe then functioned as a signal primer to initiate the dendritic rolling circle amplification reaction, containing a circular DNA and a structurally tailored hairpin structure. Subsequently, the circular nucleic acid template produced a complementary sequence to the Mg 2+ -dependent DNAzyme and a sequence identical to the loop region of the co-added hairpin structure. At last, a fluorescence readout signal was afforded by the DNAzyme-catalyzed cleavage of a fluorophore/quencher-modified substrate. This method enabled the analysis of the target MTase with a detection limit up to 0.36 U mL −1 , and a dynamic range was obtained from 1.0 to 10 U mL −1 . Moreover, the proposed strategy was successfully applied in real sample assay. With this assay, the inhibitors of MTase were evaluated and screened which might be helpful for the discovery of anticancer drugs. [ABSTRACT FROM AUTHOR]

In this work, personal glucose meter (PGM) as a portable electrochemical device was utilized for sensitive detection of non-glucose targets: N-gene and PCB77, respectively. DNA hydrogel, which can respond to CRISPR/Cas system, was prepared for label-free encapsulating invertase. In the presence of targets, the repeated sequence for the activation of Cas12a was obtained due to the performance of RCA. Unlike "one-to-one" recognition, activated Cas12a can efficiently cleave multiple single-stranded linker DNAs on DNA hydrogels, thus releasing many invertase that can be used for PGM detection. With the amplification of RCA and CRISPR/Cas system, high detection sensitivity can be obtained even using portable PGM. The detection limits for N-gene and PCB77 were 2.6 fM and 3.2 × 10−5 μg/L, respectively, with high specificity and good practical application performance. The developed biosensor can be used for online monitoring with the merit of low cost, easy operation and can be used for various targets analysis. A label-free strategy for CRISPR/Cas12a triggered portable PGM sensor was developed for the detection of N-gene and PCB77 using DNA hydrogel encapsulating invertase. [Display omitted] • Portable personal glucose meter was used to detect non-glucose targets. • DNA hydrogel was prepared for label-free encapsulating invertase. • CRISPR/Cas system can efficiently destroy DNA hydrogel and release invertase. • The same crRNA can be used to detect different targets. [ABSTRACT FROM AUTHOR]

Abstract: Methyl-CpG (mCpG) binding domain protein 4 (MBD4) is a member of mammalian DNA glycosylase superfamily. It contains an amino-proximal methyl-CpG binding domain (MBD) and a C-terminal mismatch-specific glycosylase domain, which is an important molecule believed to be involved in maintaining of genome stability. Herein, we determined the crystal structure of C-terminal glycosylase domain of human MBD4. And the structural alignments of other helix-hairpin-helix (HhH) DNA glycosylases show that the human MBD4 glycosylase domain has the similar active site and the catalytic mechanisms as others. But the different residues in the N-terminal of domain result in the change of charge distribution on the surface of the protein, which suggest the different roles that may relate some diseases. [Copyright &y& Elsevier]