RSC Advances
·
2017
·Vol. 7
·期 22
·13177-13183
·
IF 3.108
Yunyun Huang,Bo Yu,Tuan Guo,Bai-Ou Guan
Ultrasensitive and in situ DNA detection at different pH values, ranging from 4.3 to 8.5, based on a microfiber with a graphene oxide linking layer was proposed and experimentally demonstrated. The graphene oxide coating over the silica microfiber provides a strong π–π interaction with the detected single-stranded DNA molecules, and induces an amplified surface refractive index modulation over the fiber, and finally leads to a wavelength shift of the optical interference fringe for interrogation. Using this strategy, in situ DNA measurement with a detection limit of up to 10−12 M and a linear response in the pH range from 4.3–8.5 has been achieved. Benefiting from its compact size, high sensitivity, and ease of use, together with remote operational ability, the proposed sensor opens up a multitude of opportunities for quantifying DNA in various hard-to-reach environments. It may supplement the existing DNA detection tools.
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高影响因子
MICROCHIMICA ACTA
·
2018
·Vol. 186
·期 1
·1-9
·
IF 5.705
Zhang Xingxing,Liu Qiang,Jin Yan,Li Baoxin
A fluorometric method is described for the determination of the activity of the enzyme T4 polynucleotide kinase phosphatase (T4 PNKP). A short 3′-terminus phosphorylated DNA strand is hybridized with a long DNA strand to produce a partially double-stranded DNA (dsDNA) substrate. On addition of T4 PNKP, the substrate is dephosphorylated to generate the long dsDNA, and then the long dsDNA acted as a template for synthesizing copper nanoclusters (CuNCs). The dsDNA-templated CuNCs display fluorescence with excitation/emission peak wavelengths of 340/570 nm. The fluorescence is DNA sequence-dependent. A DNA substrate was designed to enhance fluorescence and to reduce the background in order to improve the sensitivity of the assay. The assay has an analytical range that extends from 0.07 U mL −1 to 15 U mL −1 and a detection limit of 0.06 U mL −1 .
The sequence-dependent fluorescence of DNA-templated copper nanoclusters, which are in-situ synthesized through the reduction of CuSO 4 by ascorbate (Vc) in the presence of dsDNA template, is utilized to obtain the method for sensitive detection of T4 PNKP activity with near-zero background.
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高影响因子
TALANTA
·
2018
·
IF 4.244
Guirong Liu,Wenli He,Chenghui Liu
As an important base excision repair (BER) enzyme, uracil-DNA glycosylase (UDG) can repair the uracil-induced DNA lesion and maintain the genomic integrity. Herein, we have proposed a sensitive UDG assay based on the terminal deoxynucleotidyl transferase (TdT)-assisted formation of fluorescent copper nanoclusters (CuNCs). In this study, a uracil-containing stem-loop DNA substrate is rationally designed with its 3′-end blocked with 2′, 3′-dideoxycytosine (ddC). UDG can remove the uracil in the DNA substrate to generate an apurinic/apyrimidinic (AP) site, which can then be specifically cleaved by endonuclease IV (Endo IV) to expose a 3′-OH terminus. TdT will initiate the template-free DNA extension along the exposed 3′-OH terminus to produce a quite long poly(T) tail, which will perfectly template the production of fluorescent CuNCs. By recording the fluorescence of the CuNCs, the UDG activity can be faithfully detected. In contrast, if UDG is absent, the 3′-ddC terminus of the DNA substrate cannot be recognized by TdT and thus no TdT-based extension and formation of CuNCs will occur. The use of ddC as a 3′-end blocker can greatly decrease the nonspecific DNA extension and improve the signal-to-noise ratio. Furthermore, TdT is a template-free and sequence-independent DNA polymerase, which can effectively catalyze the tailing process up to a maximum of thousands of thymines , and each tail can form many fluorescent CuNCs. Therefore, an ultrahigh sensitivity is achieved and as low as 0.00005 U/mL of UDG can be clearly detected. Moreover, with an additional AP site-contained poly(A) oligonucleotide during TdT-mediated extension, a branched amplification mechanism is also preliminarily devised, which can further push the detection limit of UDG to an extremely low level of 0.000002 U/mL.
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高影响因子
BIOMACROMOLECULES
·
2025
·
IF 5.4
Lin Zhang,Jiaxing Zhang,Lvping Wu,Shengping You,Rongxin Su,Wei Qi
Single-stranded DNA-binding protein gp32 is essential for DNA replication and widely used in isothermal amplification techniques such as recombinase polymerase amplification (RPA). However, its inherent thermal instability and reaction condition sensitivity restrict its broader applications. Here we develop the Fermentation Integrated Rational Engineering (FIRE-3S) strategy to enhance the functional properties of gp32 through molecular dynamics (MD) simulations, structure-guided fermentation tuning and stability-focused rational design. Starting from a disulfide-bonded mutant (V62C/T80C) that showed enhanced RPA performance but reduced thermostability, MD simulations revealed local structural disruption near the Zn2+-binding motif. Then, structure-guided fermentation tuning was applied, which restored structural integrity and enhanced DNA-binding affinity. Building on 2S-step, a computationally guided clique-based design pipeline rapidly identified a multisite mutant, gp32-M1, with elevated thermostability (ΔTm = +6.03 °C), robust ssDNA-binding, and strong inhibitor tolerance─maintaining >65% activity under 4% isopropanol or ethanol. This work establishes a systematic framework for engineering ssDNA-binding proteins with enhanced biostability and environmental resilience for molecular diagnostics.
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高影响因子
SENSORS AND ACTUATORS B-CHEMICAL
·
2016
·
IF 4.758
Chunying Li,Chunying Wei
We have constructed a label-free fluorescent probe using ssDNA-templated silver nanoclusters (Ag NCs), and its quantum yield (QY) was significantly higher (∼29%) compared to traditional water-soluble fluorescent metal nanoclusters. The proposed approach is simple, non-toxic, sensitive, and selective for the detection of mercuric ion (Hg 2+ ). The fluorescence intensity of ssDNA-Ag NCs was quenched significantly in the presence of Hg 2+ , which might be attributed to the strong interactions between them including the dynamic and static quenching mechanisms. Though Ag + induced a moderate interference to Hg 2+ , there was a negligible quenching in the presence of other metal cations. In addition, Hg 2+ could be detected as low as 4.5 nM, within a good linear range from 0 to 150 nM ( R 2 = 0.995). More importantly, this method has also been successfully applied to the detection of Hg 2+ in the real water samples. Thus, the ssDNA-Ag NCs detection system is potentially suitable for Hg 2+ sensing in environmental samples.
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高影响因子
TALANTA
·
2017
·
IF 4.162
Dan Han,Chunying Wei
In this work, we develop a fluorescent molecular beacon based on the DNA-templated silver nanoclusters (DNA-Ag NCs). The skillfully designed molecular beacon can be conveniently used for detection of diverse virulence genes as long as the corresponding recognition sequences are embedded. Importantly, the constructed detection system allows simultaneous detection of multiple nucleic acids , which is attributed to non-overlapping emission spectra of the as-synthesized silver nanoclusters. Based on the target-induced fluorescence enhancement, three infectious disease-related genes HIV, H1N1, and H5N1 are detected, and the corresponding detection limits are 3.53, 0.12 and 3.95 nM, respectively. This design allows specific, versatile and simultaneous detection of diverse targets with easy operation and low cost.
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高影响因子
ACS Applied Materials & Interfaces
·
2020
·
IF 8.758
Zhenqiang Fan,Zongqiong Lin,Zepeng Wang,Jianfeng Wang,Minhao Xie,Jianfeng Zhao,Kai Zhang,Wei Huang
In this work, we fabricated a dual-wavelength electrochemiluminescence ratiometric biosensor based on electrochemiluminescent resonance energy transfer (ECL-RET). In this biosensor, Au nanoparticle-loaded graphitic phase carbon nitride (Au-g-C3N4) as a donor and Au-modified dimethylthiodiaminoterephthalate (TAT) analogue (Au@TAT) as an acceptor were investigated for the first time. Besides, tetrahedron DNA probe was immobilized onto Au-g-C3N4 to improve the binding efficiency of the transcription factor and ECL ratiometric changes on the basis of the ratio of ECL intensities at 595 and 460 nm, which were obtained through the formation of a sandwich structure of DNA probe–antigen–antibody. Our biosensor achieved the assay of NF-κB p50 with a detection limit of 5.8 pM as well as high stability and specificity.
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高影响因子
SENSORS AND ACTUATORS B-CHEMICAL
·
2020
·
IF 7.1
Tianjiao Meng,Huixian Jia,Siying An,Huan Wang,Xinjian Yang,Yufan Zhang
Herein, a Pd nanoparticles-DNA layered nanoreticulation (Pd NPs-DNA LNR) sensor was assembled to a graphene oxide (GO)-modified electrode by catalytic hairpin assembly (CHA) and a self-assembly process. In this process, GO was modified on the electrode surface to provide -COOH sites for DNA bonding. Moreover, with the CHA circular amplification strategy, many single DNA1 were exposed to be able to be bound with Pd-DNA2 for the formation of the Pd NPs-DNA LNR, which enabled the ultrasensitive detection of microRNA-21 (miR-21) with a low limit of detection 63.1 aM. This strategy combines the amplification technology of CHA and Pd NPs-DNA LNR with Pd NPs. Such a layered nanoreticulation material can support abundant Pd NPs to avoid aggregation and obtain enhanced electrochemical signals. Significantly, this method can be successfully applied in tumor cell lysates and provides a neoteric channel for early disease diagnosis.
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高影响因子
PESTICIDE BIOCHEMISTRY AND PHYSIOLOGY
·
2019
·
IF 2.87
Benshui Shu,Jingjing Zhang,Zhiyan Jiang,Gaofeng Cui,Sethuraman Veeran,Guohua Zhong
Harmine , a useful botanical compound, has demonstrated insecticidal activity against some pests. However, harmine's mechanism of action has not been thoroughly elucidated to date. To preliminarily explore harmine's insecticidal mechanisms, the cytotoxicity of harmine against Spodoptera frugiperda Sf9 cells was comprehensively investigated. Our results indicated that harmine induced apoptosis in Sf9 cells, as evidenced by cellular and nuclear morphological changes, DNA laddering and increases in caspase-3-like activities. In addition, activation of the mitochondrial apoptotic pathway by harmine was confirmed by the generation of ROS, opening of mitochondrial permeability transition pores (MPTPs), increase in cytosolic Ca 2+ , changes in mRNA expression levels of genes involved in the mitochondrial apoptotic pathway and increase and release of Cytochrome c. Furthermore, lysosomal membrane permeabilization, release of cathepsin L from the lysosome into the cytosol and cleavage of caspase-3 were also triggered, which indicated that lysosomes were involved in this physiological process . Moreover, the effect of harmine on DNA topoisomerase I activity was investigated by in vivo and molecular docking experiments. These data not only verified that harmine induced apoptosis via comprehensive activation of the mitochondrial and lysosomal pathways and inhibition of DNA topoisomerase I activity in Sf9 cells but also revealed a mechanism of harmine insecticidal functions for pest control.
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高影响因子
JOURNAL OF PHOTOCHEMISTRY AND PHOTOBIOLOGY A-CHEMISTRY
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2014
·
IF 2.291
Li-Na Wen,Meng-Xia Xie
s G-quadruplex DNA is an effective target for anticancer drug, and investigation of the G-quadruplex DNA with natural compounds plays a key role in the development of anticancer drugs. In this article, room temperature phosphorescence sensing system based on l -cysteine-capped Mn-doped ZnS QDs has been established. Sanguinarine can adsorb on the surface of Mn-doped QDs and quenched the phosphorescence emission of the sensor. When the G-quadruplex sequences combined with the sanguinarine of the QDs-sanguinarine system, the phosphorescence intensities of the QDs sensors would be restored, and the binding abilities of the G-quadruplex DNA with the sanguinarine can be evaluated by the phosphorescence recoveries of the sensor system. Based on the developed approach, the interaction of sanguinarine and six G-quadruplex DNA with various sequences (tel24-1, tel24-2, tel22, k-ras, HIF-1α and c-kit1) has been investigated. The results showed that the binding abilities of the G-quadruplex DNA with sanguinarine were related to their number of bases and conformations. The conformational changes of the six G-quadruplex DNA, after binding with sanguinarine, have also been probed by circular dichroism. Combining the changes of the phosphorescence lifetime, the binding mechanism of the G-quadruplex DNA and sanguinarine has been discussed. The developed method was simple, rapid and specific, and can avoid fluorescence interferences of the system, and it would be prospective to evaluate the interaction of the drug with different G-quadruplexes.
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BIOMATERIALS
·
2017
·
IF 8.402
Si-An Xie,Tao Zhang,Jin Wang,Feng Zhao,Yun-Peng Zhang,Wei-Juan Yao,Sung Sik Hur,Yi-Ting Yeh,Wei Pang,Li-Sha Zheng,Yu-Bo Fan,Wei Kong,Xian Wang,Jeng-Jiann Chiu,Jing Zhou
Cells perceive the physical cues such as perturbations of extracellular matrix (ECM) stiffness, and translate these stimuli into biochemical signals controlling various aspects of cell behavior, which contribute to the physiological and pathological processes of multiple organs. In this study, we tested the hypothesis that during arterial stiffening , vascular smooth muscle cells (SMCs) sense the increase of ECM stiffness, which modulates the cellular phenotype through the regulation in DNA methyltransferases 1 (DNMT1) expression. Moreover, we hypothesized that the mechanisms involve intrinsic stiffening and deficiency in contractility of vascular SMCs. Substrate stiffening was mimicked in vitro with polyacrylamide gels. A contractile-to-synthetic phenotypic transition was induced by substrate stiffening in vascular SMCs through the down-regulation of DNMT1 expression. DNMT1 repression was also observed in the tunica media of mice aortas in an acute aortic injury model and a chronic kidney failure model, as well as in the tunica intima of human carotid arteries with calcified atherosclerotic lesions. DNMT1 inhibition facilitates arterial stiffening in vivo and promotes osteogenic transdifferentiation , calcification and cellular stiffening of vascular SMCs in vitro . These effects may be attributable, at least in part, to the role of DNMT1 in regulating the promoter activities of Transgelin (SM22α) and α-smooth muscle actin (SMA) and the functional contractility of SMCs. We conclude that DNMT1 is a critical regulator that negatively regulates arterial stiffening via maintaining the contractile phenotype of vascular SMCs. This research may facilitate elucidation of the complex crosstalk between vascular SMCs and their surrounding matrix in healthy and in pathological conditions and provide new insights into the implications for potential targeting of the phenotypic regulatory mechanisms in material-related therapeutic applications.
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高影响因子
SENSORS AND ACTUATORS B-CHEMICAL
·
2014
·
IF 3.84
Yizhong Shen,Shaopu Liu,Jidong Yang,Linlin Wang,Xuanping Tan,Youqiu He
A novel and sensitive turn-on fluorescent biosensor for herring sperm DNA (hsDNA) detecting was designed based on hsDNA-induced fluorescence enhancement of Sm 3+ -modulated glutathione (GSH)-capped CdTe quantum dots (QDs). The fluorescent biosensor was based on the following two properties. First, the high fluorescence of GSH-capped CdTe QDs could be effectively quenched by Sm 3+ due to the binding of Sm 3+ to the GSH on the surface of the QDs and the electron transfer from the photoexcited GSH-capped CdTe QDs to Sm 3+ . Second, the high affinity of hsDNA to Sm 3+ enabled Sm 3+ to be dissociated from the surface of GSH-capped CdTe QDs to form more stable complex with hsDNA in aqueous solution, thereby enhancing the fluorescence of Sm 3+ -modulated GSH-capped CdTe QDs. Experimental results showed that the enhanced fluorescence intensity of Sm 3+ -modulated GSH-capped CdTe QDs was directly proportional to the concentration of hsDNA in the range from 0.012 to 14.0 μg mL −1 with a good correlation coefficient of 0.9977, and the detection limit (3 σ / S ) was 3.61 ng mL −1 . To further investigate perfect analysis performance, the developed biosensor was applied to the determination of hsDNA in synthetic samples and human fresh serum samples with satisfactory results.
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高影响因子
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