Nature Protocols
·
2024
·
IF 13.1
Chen Feng,Li Xinyin,Bai Min,Zhao Yongxi
Epigenetic modifications and spatial proximities of nucleic acids and proteins play important roles in regulating physiological processes and disease progression. Currently available cell imaging methods, such as fluorescence in situ hybridization (FISH) and immunofluorescence, struggle to detect low-abundance modifications and their spatial proximities. Here we describe a step-by-step protocol for three DNA-encoded amplifying FISH-based imaging strategies to overcome these challenges for varying applications: base-encoded amplifying FISH (BEA-FISH), pairwise proximity-differentiated amplifying FISH (PPDA-FISH) and cellular macromolecules-tethered DNA walking indexing (Cell-TALKING). They all use the similar core principle of DNA-encoded amplification, which transforms different nonsequence molecular features into unique DNA barcodes for in situ rolling circle amplification and FISH analysis. This involves three key reactions in fixed cell samples: target labeling, DNA encoding and rolling circle amplification imaging. Using this protocol, these three imaging strategies achieve in situ counting of low-abundance modifications alone, the pairwise proximity-differentiated visualization of two modifications and the exploration of multiple modifications around one protein (one-to-many proximity), respectively. Low-abundance modifications, including 5-hydroxymethylcytosine, 5-formylcytosine, 5-hydroxymethyluracil and 5-formyluracil, are clearly visualized in single cells. Various combinatorial patterns of nucleic acid modifications and/or histone modifications are found. The whole protocol takes ~2–4 d to complete, depending on different imaging applications.
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PubMed
高影响因子
RSC Advances
·
2024
·Vol. 14
·期 38
·28077-28085
·
IF 3.9
Xiaoli Qi,Hui Hu,Lina Liang,Yuqing Lin,Yudan Liu,Haifeng Sun,Yunxian Piao
Bagasse-derived biochar nanoparticles obtained under a low pyrolysis condition (400 °C) were first revealed to be capable of highly efficiently quenching the fluorescence of 6-carboxyfluorescein, with a significantly improved quenching rate constant over that of other quenchers and high-temperature prepared ones, and were designated as bagasse-derived quencher nanoparticles (BQNPs). The BQNPs are suitable for the construction of fluorescence nanoprobes, taking advantage of their various beneficial properties, including low cost, environmental friendliness, high dispersibility, and rich functional groups that allow their easy and versatile molecular modification. They were demonstrated to be capable of stably binding single-stranded oligonucleotides through both adsorption and covalent interactions and were utilized for the construction of both BQNPs/DNA and BQNPs/aptamer probes. The BQNPs/DNA probe had strong resistance against degradation by deoxyribonuclease I and showed high precision and selectivity for the detection of single-stranded DNA, with a limit of detection of 1.04 nM. Moreover, the BQNPs/aptamer probe demonstrated the rapid and sensitive detection of 17β-estradiol (E2) with a limit of detection of 0.4 ng mL−1 with no cross-reactivity with the analogues, and it was also applied for real environmental sample detection and demonstrated reasonable signal recoveries. Benefiting from their strong quenching ability, low cost, and great dispersibility, the BQNPs show great potential for the development of cost-effective and sensitive fluorescence sensors.
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高影响因子
ANALYTICA CHIMICA ACTA
·
2019
·
IF 5.256
Zhihong Li,Hongjing Sun,Xinyue Ma,Ruifang Su,Rui Sun,Chuanyu Yang,Chunyan Sun
A novel label-free fluorescence “turn-on” strategy was developed for the sensitive detection of Hg 2+ based on the thymine-Hg 2+ -thymine (T-Hg 2+ -T) coordination and the fact that single-stranded DNA (ssDNA) greatly enhances the fluorescence of terbium (III) (Tb 3+ ), but double-stranded DNA (dsDNA) does not. In the absence of Hg 2+ , the mercury-specific DNA (MSD) hybridized with the corresponding complementary strand (cDNA) to form a double helix structure in solution based on Watson-Crick base pairings, which cannot enhance the fluorescence of Tb 3+ . In the presence of Hg 2+ , MSD preferentially bound with Hg 2+ to form the T-Hg 2+ -T complex due to the strong affinity of Hg(II) for the T bases of DNA, thus avoiding the hybridization of the cDNA to MSD. The free cDNA can greatly enhance the emission of Tb 3+ , leading to a sharp increase in fluorescence intensity . Under the optimized conditions, the increased fluorescence intensity was proportional to the concentration of Hg 2+ in the range from 10 to 600 nM, and this method can detect concentrations of Hg 2+ as low as 0.24 nM. Moreover, satisfactory results were obtained for the detection of Hg 2+ in river water and fish samples, and the results were consistent with those from the atomic fluorescence spectroscopy (AFS). Thus, the fluorescence characteristics of Tb 3+ were here used in a “turn on” approach to detect Hg 2+ , which represents a new opportunity for Hg 2+ analysis in the field of environmental monitoring and food safety.
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BIOSENSORS & BIOELECTRONICS
·
2022
·
IF 12.545
Chang Liu,Siyu Gou,Yanhui Bi,Qi Gao,Juanjuan Sun,Shanjin Hu,Weiwei Guo
A portable, cost-effective and storable DNA-gold nanoparticle (AuNP) hybrid hydrogel film based biosensing system was developed, with AuNPs serving as both the crosslinking units of the film and the signaling units. Using a layer-by-layer assembly method, hydrogel film composed of three-dimensional hydrophilic network of densely packed AuNPs interconnected by responsive DNA structures was constructed onto a glass slide. By programming the sequence of DNA structures, target-responsive hybrid films were constructed. As a proof of concept , the sequence of a substrate DNA which can be identified and cleaved by Pb 2+ -dependent DNAzyme was encoded to construct Pb 2+ -responsive DNA-AuNP hybrid hydrogel film. The high-density packing of AuNPs as signal substances significantly improved the sensitivity of the ultrathin film biosensing system while reduced the cost of expensive DNA materials. A hydrogel film composed of 10 layers of assembled DNA-AuNP structures generated sufficient visual colorimetric signals for Pb 2+ detection, with a detection limit of 2.6 nM. By introducing UO 2 2+ -dependent DNAzyme, the system could be further applied in the sensitive and selective detection of UO 2 2+ , with a detection limit of 10.3 nM. Compared with bulk-sized DNA hydrogel biosensing systems, the DNA-AuNP hydrogel film biosensing system exhibited faster response thanks to the sub-micrometer ultrathin film structures. Moreover, the protection of fragile non-covalently crosslinked DNA films with solid slides also facilitated the portable application and long-term storage of the resulting biosensing system, with 95% of the response signal retained after three months of storage. The DNA-AuNPs hydrogel film biosensing system is highly promising for future rapid on-site detection applications.
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高影响因子
BIOSENSORS & BIOELECTRONICS
·
2015
·
IF 6.409
Qiang Chen,Junfeng Zuo,Jinfeng Chen,Ping Tong,Xiujuan Mo,Lan Zhang,Jianrong Li
In this paper, a novel label-free fluorescent biosensor for terbium (ш) (Tb 3+ ) was proposed based on structural conversion of G-quadruplex DNA mediated by Thioflavin T (ThT) and Tb 3+ . In the presence of K + , ThT could bind to K + -stabilized parallel G-quadruplex, giving rise to high fluorescence intensity . Upon the addition of Tb 3+ , Tb 3+ could competitively bind to parallel G-quadruplex leading to the structural change, which resulted in fluorescence decrease. The change of fluorescence intensity (Δ F = F 0 – F ) showed a good linear response toward the concentration of Tb 3+ over the range from 1.0 pM to 10.0 µM with a limit of detection of 0.55 pM. This proposed biosensor was simple and cost-effective in design and in operation with ultrahigh sensitivity and selectivity . Thus, the proposed biosensor could be a promising candidate for monitoring ultratrace Tb 3+ in environment.
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高影响因子
JOURNAL OF SOILS AND SEDIMENTS
·
2015
·Vol. 16
·期 3
·954-965
·
IF 2.139
Ye Mao,Sun Mingming,Wan Jinzhong,Zhao Yu,Xie Shanni,Tian Da,Hu Feng,Li Huixin,Zong Lianggang,Kengara Fredrick Orori,Xin Jiang
Purpose Remediation of polybrominated diphenyl ethers (PBDEs)/heavy metals/antibiotics mixed contaminated soil with high abundance of antibiotic resistance genes (ARGs) has being greatly challenging. Materials and methods An elevated soil washing procedure was developed to investigate the potential of applying DNA solution to extract PBDEs, Cu, Pb, tetracycline, and sulfadiazine from a high abundance of ARG-affected soil, followed by microbial augmentation to restore the microbiological functioning of washed soil. Results and discussion Two successive washings with 15 g L −1 DNA solution plus ultrasonication (25 kHz for 15 min) were optimal for extracting 80.6 % BDE28, 85.2 % BDE47, 64.7 % Cu, 60.3 % Pb, 100 % tetracycline, and 100 % sulfadiazine. Simultaneously, the ARG abundance decreased to the level of 10 −7 –10 −8 (ARGs copies/16S copies). Moreover, combined treatment with PBDE-degrading bacteria inoculation and nutrient application for 3 months to the second washed soil resulted in the further degradation of residual BDE28 and BDE47. Furthermore, the bacterial community composition was restored and the transfer risk of residual pollutants was limited, as indicated by high-throughput sequencing technology analysis and heavy metals stability model evaluation. Conclusions The combined cleanup technology is an environmentally friendly and effective technology to remediate mixed contaminated soils.
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Polymer Chemistry
·
2014
·Vol. 6
·期 5
·780-796
·
IF 5.368
Cheng Wang,Xiuli Bao,Xuefang Ding,Yang Ding,Sarra Abbad,Yazhe Wang,Min Li,Yujie Su,Wei Wang,Jianping Zhou
A new pH-responsive charge-conversion and rapidly self-dissociative multifunctional polymer lauric acid-polyethyleneimine-cyclohexanedicarboxylic anhydride-folic acid (LA-PEI-HHPA-FA, LPHF) is developed. LPHF as an amide with neighboring carboxylic acid groups becomes negatively charged at physiological pH and was used as a coating polymer to shield positively charged DNA/polyethyleneimine 25 k (DP) binary polyplexes, resulting in the formation of DPLPHF ternary polyplexes. The characterization using various methods proved that DPLPHF ternary polyplexes are stable at physiological pH, while in acidic endosomes, the coating polymer LPHF could quickly self-dissociate from the DP binary polyplexes. As compared with DP binary polyplexes, DPLPHF ternary polyplexes containing the tumor targeting ligand folic acid (FA), the absorption enhancer lauric acid (LA) and the charge-conversion entity cis-1,2-cyclohexanedicarboxylic anhydride (HHPA) exhibited more effective cellular uptake and higher transfection efficiency with lower non-therapeutic cytotoxicity in vitro. Moreover, in vivo studies revealed the significantly stronger tumor targeting and tumor inhibiting efficacy of DPLPHF ternary polyplexes. All these findings demonstrated the potential of LPHF as a promising coating polymer for enhanced gene transfection efficiency of DP binary polyplexes in vitro and in vivo.
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高影响因子
Science of Advanced Materials
·
2022
·Vol. 14
·期 5
·911-919
·
IF 1.067
Zhu Chunxia,Ming Shenqiu,Qiu Bin,Zhong Hui
FePt nanoparticles were in this study prepared at varied concentrations to assess their mechanism in regulating DNA injury and inactivation of Hela cells through mitogen-activated protein kinase (MAPK) signal pathway. The cells were divided into following 5 group sets: 1st, 2nd, 3rd,
4th and 5th, which were randomly intervened by following varied concentrations of FePt nanoparticles; 0, 5, 10, 15 and 20 μg/mL. Their action on cellular toxicity, proliferation and inactivation of cells was detected with CCK-8 method, while apoptotic condition was detected with
flow cytometry (FCM). The degree of DNA double chains fracture was detected with neutral single cell gel electrophoresis and immunofluorescence staining. The protein expressions of of EKR1/2, p-ERK1/2, p38 MAPK and p-p38 MAPK in every set were detected with Western Blot assay, while proliferative
activity in the 4th and 5th sets was lower than in the 1st set, and inhibiting action in the 4th set was best. There was no notable difference between 4th and 5th sets, and apoptotic rate in the 1st set was lower than in the other sets. The DNA percentage in the Hela cells was reduced and
Tial length was lengthened along with increased concentration of adopted FePt nanoparticles. The proliferation of Hela cells was restrained by FePt nanoparticles through restraining of the MAPK signal pathway, but growth of Hela cells was not affected, and DNA double-strand break (DSB) was
increased. The FePt nanoparticles could therefore be adopted as one kind of effective anti-tumor drug through increased DNA DSB in tumor cells based on killing effect of tumor cells and good compatibility for normal cells.
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DALTON TRANSACTIONS
·
2020
·Vol. 49
·期 16
·5192-5204
·
IF 4.174
Yan Chen,Qingpeng Wang,Zuojie Li,Zhifang Liu,Yanna Zhao,Junfeng Zhang,Min Liu,Zhengping Wang,Dacheng Li,Jun Han
Cycloxygenases (COXs) and matrix metalloproteinases (MMPs) in the tumor microenvironment (TME) are tightly related to the progression of cancers. Here, naproxen as a potent inhibitor of both COX and MMP was combined with platinum(IV) to construct hybrids as antitumor agents. Compound 2 with comparable or even superior activities to that of cisplatin, oxaliplatin, and carboplatin, great potential for reversing drug resistance, and superior tumor targeting properties was screened out as a lead compound. Moreover, compound 2 possessed potent tumor growth inhibition capability in vivo, which was comparable to that of oxaliplatin, and displayed rather lower side effects than the platinum(II) reference drugs. The naproxen platinum(IV) complex could easily undergo reduction and liberate the platinum(II) complex and naproxen as well as exert a multifunctional antitumor mechanism: (i) the liberated platinum(II) fragment would cause serious DNA injury; (ii) naproxen would inhibit COX-2 and decrease tumor-associated inflammation; and (iii) the naproxen platinum(IV) complex exhibited remarkable MMP-9 inhibition in tumor tissues. These antitumor functions can help reduce the growth and metastasis of malignancy.
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高影响因子
SENSORS AND ACTUATORS B-CHEMICAL
·
2020
·
IF 7.1
Bo-Xin Zheng,Wei Long,Yi-Han Zhang,Xuan-He Huang,Cui-Cui Chen,Dong-Xiao Zhong,Meng-Ting She,Ze-Xin Chen,Dong-Peng Cai,Yu-Jing Lu,Wing-Leung Wong
Red fluorescent G-quadruplex DNA selective organic binding probe is relatively underdeveloped. In the present study, we designed a series of new fluorescent probes that exhibit large Stokes shift (112−131 nm), high quantum yield (up to 0.56), robust against photo-bleaching and high selectivity towards quadruplex structures of c-MYC and telomere DNA with strong binding affinity . The discrimination ability of the probes towards G-quadruplex structures against single-/double-stranded DNA or RNA was investigated by analyzing their structural property with respect to their binding specificity. From the sensing application of the probe targeting pu27, an excellent linear relationship (R 2 = 0.9997) with respect to the pu27- A3 interaction signal was observed. The LOD was found down to 4.0 nM, which is superior to thiazole orange dye. The probe is able to realize real-time fluorescence visualization in aqueous, PAGE staining and live cell imaging. In addition, from the immunofluorescence analysis using the probe and BG4 antibody, the A3 foci and BG4 staining co-localized well in PC3 cells, which suggest that the probe is G-quadruplex selective and is useful for visualizing G-quadruplex DNA structures in live cells. Cytotoxicity assays also demonstrated that the probe has low cytotoxicity against both PC3 cancer cells and 16HBE normal cells.
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高影响因子
JOURNAL OF COORDINATION CHEMISTRY
·
2024
·
IF 2.2
Ying Zhou,Xiao-Tong Song,Hui-Chao Guan,Shu-Mei Yue
In this investigation, six new metal complexes with 2-(pyridin-2-yl)benzo[d]imidazole (bpm) were synthesized and analyzed by single-crystal X-ray diffraction (XRD), including catena-bis[(µ-2,2-bis(4-carobxyphenyl)hexafluoropropane)-(2-(pyidin-2-yl)-1H-benzimidazole)-aqua-zinc] (1), catena-[(µ-2,2-bis(4-carobxyphenyl)hexafluoropropane)-(2-(pyidin-2-yl)-1H-benzimidazole)-aqua-cadmium] (2), aqua-[2,2-bis(4-carobxyphenyl)hexafluoropropane]-bis[2-(pyridin-2-yl)-1H-benzimidazole]-manganese (3), μ-[2-nitrobenzene-1,4-dicarboxylato]-μ-[2-nitrobenzene-1,4-dicarboxylato]-bis[2-(pyridin-2-yl)-1H-benzimidazole]-di-zinc (4), catena-[bis(µ-2-nitrobenzene-1,4-dicarboxylato)-diaqua-bis(2-(pyridin-2-yl)-1H-benzimidazole)-di-cadmium] (5), aqua-[pyridine-2,6-dicarboxylato]-[2-(pyridin-2-yl)-1H-benzimidazole]-copper hydrate (6) (H2hfipbb = 2,2-bis(4-carboxyphenyl)hexafluoropropane, H2nbdc acid = Nitroterephthalic acid, and H2pda = 2,6-pyridinedicarboxylic acid). Complex 1 crystallized in a monoclinic system with space group P21/n; 2 crystallized in a monoclinic system with space group C2/c; 3 crystallized in an orthorhombic system with space group Pna21; 5 crystallized in a monoclinic system with space group P21/c; 4 and 6 crystallized in triclinic systems with space group P-1. The DNA-binding modes of the complexes were investigated by several analysis methods (e.g. viscosity analysis, circular dichroism spectroscopy, fluorescent spectroscopic analysis, as well as UV spectroscopic analysis). Complexes 1-6 bind to calf thymus DNA via an intercalative mode. The results of this study provide insights into the mechanism of DNA binding and provide a basis for rationally designing new powerful reagents for DNA detection and targeting.
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CHEMICAL ENGINEERING JOURNAL
·
2026
·
IF 12.5
Tingting Yu,Peng Song,Ruixue Wang,Ziwen Chen,Liang Zhao,Ming Wang,Jianying Wang,Tao Mei,Xianbao Wang,Jinhua Li
Colorectal cancer (CRC) is one of the malignant tumors with high morbidity and mortality rates worldwide. Conventional screening methods, such as endoscopy and fecal testing, are invasive and have low compliance, highlighting the urgent need for efficient, non-invasive diagnostics. Circulating small RNAs, including the sequence annotated as PIWI-interacting RNA-54265 (piR-54265), have been reported to be closely associated with CRC and represent promising serum biomarkers in liquid biopsy research. This study aims to develop an ultrasensitive, label-free solution-gated graphene transistor (SGGT) based biosensor to detect piR-54265 in serum samples, addressing limitations of current CRC biomarker detection. The Polyadenine 15 -thymine 5 (PolyA 15 T 5 -DNA) probe was designed with an L-shaped configuration, constructing a sensing interface. This configuration is proposed to facilitate a favorable probe conformation for nucleic acid recognition at the gate interface, thereby contributing to the overall sensing performance of the biosensor. The biosensor achieves a remarkable limit of detection (LOD) of 8.56 × 10 −19 M, with a linear detection range from 1 aM to 10 nM, and excellent selectivity for piR-54265. Clinical serum samples analyzed under diluted conditions demonstrate clear differentiation between CRC patients and non-cancer controls, with receiver operating characteristic (ROC) analysis further confirming its exceptional diagnostic performance and achieving an area under the curve (AUC) of 1.0. These results establish a SGGT-based sensing platform for CRC-associated RNA detection and highlight its potential for non-invasive biomarker detection and future diagnostic applications.
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