ANALYTICAL SCIENCES
·
2017
·Vol. 33
·期 10
·1155-1160
·
IF 1.228
He Chenmeng,Tang Yali,Wang Sai,Liu Jiahui,Chen Ying,Dong Yiyang,Su Haijia,Tan Tianwei
A simple and reliable electrochemical biosensor based on an in situ synthetic Au-reduced graphene oxide (Au-RGO) nanocomposite was constructed in this work, which is considered to be a potential sensing platform for sensitive and selective Bt63 rice detection. UV-vis spectroscopy, transmission electron microscopy (TEM), and cyclic voltammograms of the surface characterization indicated that RGO was prepared successfully and Au nanoparticles were well dispersed on its surface with an average size of 20 nm. The square-wave voltammetry (SWV) response of the electrochemical marker methylene blue (MB) was chosen to monitor the probe immobilization and target hybridization event. Under the optimum conditions, the peak values of MB were linear with the logarithm of the target DNA concentrations being from 1.0 × 10 –9 to 1.0 × 10 –14 M; the detection limit was 3.36 × 10 –15 M. Studies also demonstrated that the DNA biosensor had high reproducibility and stability. At last, we used this biosensor to detect several rice samples; it showed good sensitivity and selectivity. Thus the applicability of the method as an effective tool for genetically modified organism (GMO) quantification was confirmed by its accurate and sensitive results in Bt63 rice screening.
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ANALYTICAL CHEMISTRY
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2022
·
IF 8.008
Minghua Deng,Jinhua Li,Bichen Xiao,Zhanpeng Ren,Ziqin Li,Haiyang Yu,Jiashen Li,Jianying Wang,Zhaowei Chen,Xianbao Wang
Developing highly sensitive, reliable, cost-effective label-free DNA biosensors is challenging with traditional fluorescence, electrochemical, and other techniques. Most conventional methods require labeling fluorescence, enzymes, or other complex modification. Herein, we fabricate carbon quantum dot (CQD)-functionalized solution-gated graphene transistors for highly sensitive label-free DNA detection. The CQDs are immobilized on the surface of the gate electrode through mercaptoacetic acid with the thiol group. A single-stranded DNA (ssDNA) probe is immobilized on CQDs by strong π–π interactions. The ssDNA probe can hybridize with the ssDNA target and form double-stranded DNA, which led to a shift of Dirac voltage and the channel current response. The limit of detection can reach 1 aM which is 2–5 orders of magnitude lower than those of other methods reported previously. The sensor also exhibits a good linear range from 1 aM to 0.1 nM and has good specificity. It can effectively distinguish one-base mismatched target DNA. The response time is about 326 s for the 1 aM target DNA molecules. This work provides good perspectives on the applications in biosensors.
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ANALYST
·
2020
·Vol. 145
·期 12
·4254-4259
·
IF 3.978
Qingqing Zhang,Tong Yang,Guoxiang Zheng,Heng Gao,Chenxiao Yan,Xiong Zheng,Xiaoshun Zhou,Yong Shao
G-triplex (G3) has been recognized as a popular intermediate during the folding of G-quadruplex (G4). This has raised interest to anticipate the ultimate formation of G3 by shortening the G4-forming oligonucleotides with the remaining three G-tracts. Some G3 structures have been validated and their stability has been found to be affected by the loop sequences similar to G4s. In this work, however, we first found that an intermolecular parallel G4 structure was preferred in K+ for the oligonucleotide 5′-TGGGTAGGGCGGG-3′ (DZ3) containing only three G-tracts. We screened auramine O (AO) as the appropriate fluorophore with a molecular rotor feature to target this G4 structure. AO bound with DZ3 in a 1 : 4 ratio, as confirmed by isothermal titration calorimetry experiments, suggesting the formation of a tetramolecular G4 structure (4erG4). The excimer emission from the labelled pyrene and the DNA melting behavior at various pHs in the presence of Ag+ proved the formation of the 4erG4 structure rather than the prevalent intramolecular G3 folding. This work demonstrates that one should be cautious while putatively predicting a G3 structure from an oligonucleotide containing three G-tracts.
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Synthetic and Systems Biotechnology
·
2019
·
IF 0
Jingjun Mo,Shuwen Wang,Wan Zhang,Chunyu Li,Zixin Deng,Lixin Zhang,Xudong Qu
Actinobacteria able to produce varieties of bioactive natural products have been long appreciated by the field of drug discovery and development. Recently, a few of CRISPR/Cas9 systems bearing different types of replicons (pSG5 and pIJ101) were developed to efficiently edit their genomes. Despite wide application in gene editing, their utility in editing challenging DNA regions e.g. high sequence identity has not been compared. In this study, we confirmed that the widely used temperature-sensitive pSG5 replicon is indeed not suitable for editing modular polyketide synthase (PKS) genes due to causing unpredicted gene recombination. This problem can be addressed by replacing the pSG5 with the segregationally unstable pIJ101 replicon. By introducing a counter-selection marker CodA, convenient cloning sites in the single guide RNAs (sgRNAs) and homologous template scaffolds, we developed a new CRISPR-Cas9 system pMWCas9. This system was successfully used to delete/replace erythromycin PKS and other biosynthetic genes in Saccharopolyspora erythraea and Streptomyces sp. AL2110. By swapping the promoters of antB and antC with ermE and kasOp , we achieved a deacyl-antimycin hyper producer which produces a 9-fold higher yield than the original Streptomyces sp. AL2110 strain. Our results provide a robust and useful Cas9 tool for genetic studies in Actinobacteria .
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Scientific Reports
·
2013
·Vol. 3
·期 1
·1-8
·
IF 2.927
Wang Chong,Wu Congyu,Zhou Xuejiao,Han Ting,Xin Xiaozhen,Wu Jiaying,Zhang Jingyan,Guo Shouwu
Graphene quantum dots (GQDs) maintain the intrinsic layered structural motif of graphene but with smaller lateral size and abundant periphery carboxylic groups and are more compatible with biological system, thus are promising nanomaterials for therapeutic applications. Here we show that GQDs have a superb ability in drug delivery and anti-cancer activity boost without any pre-modification due to their unique structural properties. They could efficiently deliver doxorubicin (DOX) to the nucleus through DOX/GQD conjugates, because the conjugates assume different cellular and nuclear internalization pathways comparing to free DOX. Also, the conjugates could enhance DNA cleavage activity of DOX markedly. This enhancement combining with efficient nuclear delivery improved cytotoxicity of DOX dramatically. Furthermore, the DOX/GQD conjugates could also increase the nuclear uptake and cytotoxicity of DOX to drug-resistant cancer cells indicating that the conjugates may be capable to increase chemotherapy efficacy of anti-cancer drugs that are suboptimal due to the drug resistance.
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Science Advances
·
2020
·
IF 13.117
Huiyi Liang,Yanzi Yan,Jingjiao Wu,Xiaofei Ge,Lai Wei,Lixin Liu,Yongming Chen
Cell-free DNA (cfDNA) released from damaged or dead cells combines with LL37 and is converted into an immune response activator to exacerbate psoriasis. Here, we show that cationic nanoparticles (cNPs) efficiently compete for DNA from the DNA-LL37 immunocomplex and inhibit DNA-LL37-induced cell activation. Using phenotypical images, psoriasis area and severity index scoring, histology, and immunohistochemical analysis, we demonstrate that topical application of cNPs on psoriasiform skin of a mouse model relieves psoriatic symptoms. It is noteworthy that the results were confirmed in a cynomolgus monkey model. Moreover, topically administrated cNPs showed low in vivo toxicity because of their retention in skin. Mechanistic analyses of cytokine expression in the psoriatic site, cfDNA levels in circulation and inflamed skin, skin permeation, and biodistribution of cNPs also matched the therapeutic outcomes. Therefore, we present a previously unidentified strategy of nanomedicine to treat skin inflammatory diseases, which demonstrates great potential for clinical application.
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STAR Protocols
·
2022
·
IF 0
Qianqian Fang,Zhengkang Hua,Zhonghui Lin
Summary The human MUS81-EME1&2 complexes are structure-selective endonucleases that play important roles in DNA damage repair. Here, we describe a protocol to determine the endonuclease activities of MUS81-EME1&2 complexes toward various DNA structures. We co-express MUS81 with EME1 or EME2 and purify the complexes with high purity, and determine their activities on the cleavages of 3ˊ flaps, 5ˊ flaps, nicked double-stranded DNAs, and Holliday junctions. This protocol can also be used for the determination of substrate preferences of other structure-selective endonucleases. For complete details on the use and execution of this protocol, please refer to Hua et al. (2022) .
研究领域:
分子生物学
·
细胞生物学
·
遗传学
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JOURNAL OF INORGANIC BIOCHEMISTRY
·
2021
·
IF 4.155
Yueqin Li,Changhao Qian,Yun Li,Yong Yang,Dong Lin,Xiaohui Liu,Chen Chen
Two novel Fe(III) complexes, Fe(HL 1 ) 2 Cl·1.25H 2 O ( 1 ) and Fe(HL 2 ) 2 ·Et 3 NH·H 2 O ( 2 ) (H 2 L 1 = o -vanillin benzoylhydrazone, H 3 L 2 = o -vanillin salicylhydrazone) are prepared. X-ray single crystal diffraction confirms that the hydrazone ligands can be chelated to iron centre resulting in a six-coordinate octahedral configuration. Both complexes show major intercalation effect to the herring sperm deoxyribonucleic acid (HS-DNA) with high binding constants of 2.01 × 10 4 M −1 and 2.24 × 10 4 M −1 , respectively. Molecular docking studies reveal both complexes can intercalate at the gap of DC5-DG2 and DG6-DC1 base pairs of DNA hexamer (1Z3F). The interaction of the complex 1 with plasmid pBR322 DNA induces distinguishable alterations of the DNA morphology. Further, the structure of plasmid pBR322 DNA treated with complex 1 in the presence of ascorbic acid has been damaged probably due to the reactive oxygen species (ROS) generation. What's more, both complexes show high affinity with bovine serum albumin (BSA), the binding constants measured by fluorescence techniques are 5.75 × 10 6 M −1 and 4.39 × 10 7 M −1 , respectively. Molecular docking demonstrates that the complexes prefer the binding pocket of site III (subdomain IIB) of BSA (PDB ID: 4F5S ). Similarly, dynamic light scattering (DLS) reveals that the complexes not only bind to BSA but also induce bigger size aggregates as the concentration increases.
研究领域:
无机化学
·
生物化学
·
药物化学
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JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY
·
2023
·
IF 6.1
Yu-gang Shi,Wen-xuan Chen,Mei-zhi Zheng,Yue-xin Zhao,Yi-ran Wang,Yen-Ho Chu,Shao-ting Du,Ze-yu Shi,Qing Gu,Jian-she Chen
While photodynamic inactivation (PDI) has emerged as a novel sterilization strategy for drinking water treatment that recently attracted tremendous attention, its efficiency needs to be further improved. In this study, we aimed to clarify the ultraefficient mechanism by which potassium iodide (KI) potentiates octyl gallate (OG)-mediated PDI against bacteria and biofilms in water. When OG (0.15 mM) and bacteria were exposed to blue light (BL, 420 nm, 210 mW/cm2), complete sterilization (>7.5 Log cfu/mL of killing) was achieved by the addition of KI (250 mM) within only 5 min (63.9 J/cm2). In addition, at lower doses of OG (0.1 mM) with KI (100 mM), the biofilm was completely eradicated within 10 min (127.8 J/cm2). The KI-potentiated mechanism involves in situ rapid photogeneration of a multitude of reactive oxygen species, especially hydroxyl radicals (•OH), reactive iodine species, and new photocytocidal substances (quinone) by multiple photochemical pathways, which led to the destruction of cell membranes and membrane proteins, the cleavage of genomic DNA and extracellular DNA within biofilms, and the degradation of QS signaling molecules. This multitarget synergistic strategy provided new insights into the development of an environmentally friendly, safe, and ultraefficient photodynamic drinking water sterilization technology.
研究领域:
环境科学
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水处理
·
微生物学
·
光化学
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Applied Sciences-Basel
·
2021
·Vol. 11
·期 24
·11597
·
IF 2.679
Jianying Zhao,Jing Qian,Ji Luo,Mingming Huang,Wenjing Yan,Jianhao Zhang
Featured ApplicationDuring food processing, the surface of food and processing equipment is easily contaminated by microorganisms, which will cause food spoilage or the spread of disease, threatening food safety. Plasma-activated solution can be used as a novel food disinfectant, enhancing food quality. Plasma-activated solution will not leave chemical residues or cause environmental pollution. It is a green and environmentally friendly disinfectant and promises to replace traditional chemical disinfectants.AbstractPlasma-activated solution has attracted more attention in the food industry due to no chemical residue and good bacteriostatic properties. This study aimed to evaluate the effects of plasma-activated hydrogen peroxide solution (PAH) on the morphophysiology ofStaphylococcus aureusbiofilms. PAH was prepared using dielectric-barrier-discharge plasma and incubated withS. aureusbiofilms for 0–40 min. Changes in biofilm morphophysiology were evaluated with laser scanning confocal microscopy, electron microscopic images, reactive oxygen species (ROS) content, metabolic capacity, and 1% agarose gel. Results indicated that the population ofS. aureusin the biofilms was reduced by 4.04-log after incubation with PAH for 30 min. The thickness and metabolic capacity of biofilms were decreased, the ROS content and DNA fragments of bacteria increased after PAH treatments. Data suggested that PAH treatments significantly destroyed the morphophysiology ofS. aureus(ATCC 6538) biofilms and could be considered as a valuable anti-biofilm technology to reduce foodborne pathogens on food and/or in food facilities.Keywords:foodborne pathogen;cold plasma;nonthermal antimicrobial treatment;metabolic capacity;DNA damage
研究领域:
食品科学
·
微生物学
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CyTA-Journal of Food
·
2020
·
IF 1.653
Yongzhen Wang,Kai Zhu,Deguo Wang
The objective of this study was to develop a reliable visual method for rapid detection of donkey components to protect consumers from commercial adulteration. Six specific loop-mediated isothermal amplification (LAMP) primers targeted at donkey mitochondrial cytochrome b gene were designed, the mitochondrial DNA extraction simplified, the LAMP reaction system optimized, the specificity verified with mitochondrial DNA of horse, pork, cow, sheep, chicken, duck, and rabbit as negative controls, the detection limit determined with gradient dilution of adulterated meat with donkey meat, and the visual LAMP method for detection of donkey-derived ingredient in common meat products established. The results showed that the modified mitochondrial DNA extraction method was simple and repeatable, and the visual LAMP method with 4-(2-pyridylazo)-resorcinol sodium salt as indicator can accurately and specifically detect the donkey meat in common meat products, 1% detection limit. The study provided a promising solution for facilitating the surveillance of the commercial adulteration in processed meat.
研究领域:
食品科学
·
分子生物学
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MICROCHEMICAL JOURNAL
·
2022
·
IF 5.304
Meijuan Du,Qian Chen,Xueqin Xu
A novel and label-free electrochemical aptasensor based on exonuclease III (Exo III) and G-quadruplex DNAzyme has been first designed to detect metronidazole (MNZ) sensitively and selectively in this paper. Capture probe (CP), helper probe (HP) and aptamer (Apt) hybridize to form double-stranded DNA (dsDNA) structure with a blunt 3′-terminus, resulting in the guanine (G)-rich CP being enclosed in the dsDNA, followed by specific digestion of the 3′-terminus by Exo III to expose the G-rich sequence. At this time, CP can bind with the added hemin to form G-quadruplex DNAzyme which catalyzes the redox reaction of 3,3′,5,5′-tetramethylbenzidine (TMB) and H 2 O 2 , generating a strong current signal. However, in the presence of MNZ, the binding of MNZ and aptamer breaks the original blunt 3′-terminus of the dsDNA structure, blocking the subsequent series of reactions, and only a weak current signal can be observed. Under the optimal conditions, the aptasensor showed excellent sensitivity and specificity for the detection of MNZ with a wide linear range from 10 pM to 500 nM and a detection limit as low as 3.0 pM (S/N = 3). The aptasensor constructed based on MNZ aptamer can effectively avoid the interference of other nitro compounds and antibiotics to a certain extent. Compared with other specific recognition elements, the aptasensor possesses the advantages of strong specificity, simple preparation and low cost. Moreover, the proposed aptasensor has been successfully applied to the detection of MNZ in tap water and honey samples.
研究领域:
生物化学
·
分析化学
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