ANALYST
·
2012
·Vol. 138
·期 1
·197-203
·
IF 4.23
Sai Bi,Yangyang Cui,Li Li
An ultrasensitive and highly selective method for polymerase chain reaction-free (PCR-free) messenger RNA (mRNA) expression profiling is developed through a novel cross-rolling circle amplification (C-RCA) process based on DNA-rotaxane nanostructures. Two species of DNA pseudorotaxane (DPR) superstructures (DPR-I and DPR-II) are assembled by threading a linear DNA rod through a double-stranded DNA (dsDNA) ring containing two single-stranded gaps. In this assay, cDNA that is specific for β-actin (ACTB) mRNA is taken as a model analyte. Upon the introduction of the target cDNA, the cDNA and the biotin-modified primer are hybridized to the single-stranded regions of the DNA rod and the gap-ring, respectively. As a result, the DPR-I dethreads into free DNA macrocycle and a dumbbell-shaped DNA nanostructure. In the presence of DNA polymerase/dNTPs, two release-DNA on the DPR-I are replaced by polymerase with strand-displacement activity, which can act as the input of the DPR-II to trigger the dethreading of DPR-II and the RCA reaction, releasing another two specified release-DNA strands those in turn serve as the “mimic cDNA” for DPR-I. The C-RCA reaction then proceeds autonomously. To overcome the high background induced by hemin itself, the biotinylated rolling circle products are captured by streptavidin-coated MNPs, achieving a detection limit as low as 0.1 zmol cDNA. The assay also exhibits an excellent selectivity due to its unique DNA nanostructure fabricated through base pairing hybridization. The ACTB mRNA expression in mammary cancer cells (MCF-7) is successfully detected.
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BIOSENSORS & BIOELECTRONICS
·
2012
·
IF 5.602
Huanshun Yin,Yunlei Zhou,Zhenning Xu,Lijian Chen,Di Zhang,Shiyun Ai
DNA methylation is one of important epigenetics events, and responsible to transcription, genomic imprinting and cellular differentiation . Aberrant DNA methylation is always contacted with various diseases. Methyl binding domain (MBD) proteins can specifically bind to the methylated CpG dinucleotides . Conventional assay for DNA methylation normally need bisulfide treatment, methylated nucleotide labeling or PCR amplification. Here, we fabricated a novel electrochemical biosensor for detection of DNA methylation, assay of DNA methyltransferase (MTase) activity and screening of MTase inhibitor based on MBD protein and coomassie brilliant blue G250 (CBB-G250), where the electrochemical signal of CBB-G250 was used to monitor the methylation event. After the hybrids of DNA S1 and DNA S2 were treated with M. Sss I MTase in the presence of S -adenosylmethionine, the MBD proteins were specifically conjugated to the methylation site of CpG dinucleotides, and then, the MBD proteins were stained with CBB-G250. The electrochemical signal of CBB-G250 increased linearly with increasing M. Sss I MTase concentration in the range from 0.1 to 40 unit/mL. Furthermore, the inhibition investigation demonstrates that fisetin and chlorogenic acid can inhibit the M. Sss I MTase activity with the IC 50 value of 153.12 and 137.07 μM, respectively. Therefore, we think that this study may provide a sensitive platform for screening of DNA MTase inhibitors.
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ANALYTICAL BIOCHEMISTRY
·
2015
·
IF 2.219
Yanqin Yang,Shuai Zhang,Mengmeng Kang,Linghao He,Jihong Zhao,Hongzhong Zhang,Zhihong Zhang
A highly sensitive electrochemical DNA biosensor made of polyaniline (PANI) and gold nanoparticles (AuNPs) nanocomposite (AuNPs@PANI) has been used for the detection of trace concentration of Ag + . In the presence of Ag + , with the interaction of cytosine–Ag + –cytosine (C–Ag + –C), cytosine-rich DNA sequence immobilized onto the surface of AuNPs@PANI has a self-hybridization and then forms a duplex-like structure. The whole detection procedure of Ag + based on the developed biosensor was evaluated by electrochemical impedance spectroscopy . On semi-logarithmic plots of the log Ag + concentration versus peak current, the results show that the prepared biosensor can detect silver ions at a wide linear range of 0.01–100 nM ( R = 0.9828) with a detection limit of 10 pM (signal/noise = 3). Moreover, the fabricated sensor exhibits good selectivity and repeatability. The detection of Ag + was determined by Ag + self-induced conformational change of DNA scaffold that involved only one oligonucleotide , showing its convenience and availability.
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高影响因子
JOURNAL OF ELECTROANALYTICAL CHEMISTRY
·
2013
·
IF 2.672
Lixuan Zeng,Aizhen Zhang,Xiaohua Zhu,Chuyi Zhang,Yong Liang,Junmin Nan
The electrochemical determination of nonylphenol (NP) using differential pulse voltammetry (DPV) is developed based on a reduced graphene–DNA hybrid-modified glassy carbon electrode (GR-DNA/GCE). The electrochemical oxidation of NP at the GR-DNA/GCE is shown to be an one-electron and one-proton process, and due to the synergic effect of GR and DNA , the GR-DNA/GCE has 4.87, 2.76 and 2.09 times higher current response than bare GCE, DNA/GCE, and GR/GCE, respectively. Using DPV in the 0.1 mol L −1 acetate buffer solution at 0.1 V and with a 300 s accumulation time (pH = 4.6), the GR-DNA/GCE exhibits a linear current response towards the electrochemical oxidation of NP in the concentration range of 5.0 × 10 −8 to 4.0 × 10 −6 mol L −1 and a detection limit of 1.0 × 10 −8 mol L −1 ( S / N = 3). In addition, this GR-DNA/GCE also indicates high selectivity and reproducibility, and can be successfully used to determine the presence of NP in Asian Clams and natural water samples with comparative sensitivity to high-performance liquid chromatography, and with recoveries ranging from 95% to 103%, showing its practical prospects in the determination of NP in real samples.
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BIOSENSORS & BIOELECTRONICS
·
2014
·
IF 6.451
Wang Ren,Zhong Feng Gao,Nian Bing Li,Hong Qun Luo
This work reported a novel, ultrasensitive, and selective platform for electrochemical detection of DNA , employing an integration of exonuclease III (Exo-III) assisted target recycling and hybridization chain reaction (HCR) for the dual signal amplification strategy. The hairpin capture probe DNA (C-DNA) with an Exo-III 3′ overhang end was self-assembled on a gold electrode. In the presence of target DNA (T-DNA), C-DNA hybridized with the T‐DNA to form a duplex region, exposing its 5′ complementary sequence (initiator). Exo-III was applied to selectively digest duplex region from its 3-hydroxyl termini until the duplex was fully consumed, leaving the remnant initiator. The intact T-DNA spontaneously dissociated from the structure and then initiated the next hybridization process as a result of catalysis of the Exo-III. HCR event was triggered by the initiator and two hairpin helper signal probes labeled with methylene blue , facilitating the polymerization of oligonucleotides into a long nicked dsDNA molecule. The numerous exposed remnant initiators can trigger more HCR events. Because of integration of dual signal amplification and the specific HCR process reaction, the resultant sensor showed a high sensitivity for the detection of the target DNA in a linear range from 1.0 fM to 1.0 nM, and a detection limit as low as 0.2 fM. The proposed dual signal amplification strategy provides a powerful tool for detecting different sequences of target DNA by changing the sequence of capture probe and signal probes, holding a great potential for early diagnosis in gene-related diseases.
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BIOSENSORS & BIOELECTRONICS
·
2015
·
IF 6.409
Hailiang Huang,Shuo Shi,Xuyue Zheng,Tianming Yao
Based on specific homo-A/T DNA binding properties, a strategy for coralyne and mercury ions detection was realised by exonuclease-aided signal amplification . Coralyne could specifically bind homo-A DNA and protect it from the hydrolysis of exonuclease I. The coralyne-protected DNA was subsequently used as a trigger strand to hydrolyze DNA2 in exonuclease-aided signal amplification process. Thiazole orange was used to quantify the remainder DNA2. Under the optimal condition, the fluorescence intensity was linearly proportional to the concentration of coralyne in the range of 0.2–100 nM with a limit of detection (LOD) of 0.31 nM, which presented the lowest LOD for coralyne among all reported. With homo-T and Hg 2+ taking the place of homo-A DNA and coralyne, respectively, the system could also be used for Hg 2+ detection. The experiments in real samples also showed good results. This method was label-free, low-cost, easy-operating and highly repeatable for the detection of coralyne and mercury ions. It could also be extended to detect various analytes, such as other metal ions, proteins and small molecules by using appropriate aptamers.
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BIOSENSORS & BIOELECTRONICS
·
2013
·
IF 5.437
Wenjiao Zhou,Jiao Su,Yaqin Chai,Ruo Yuan,Yun Xiang
Naked eye-based detection has received increasing research interest due to the simplicity nature of this type of assay. However, improving the sensitivity of the naked eye detection method for the monitoring of trace amount of target molecules remains a major challenge. Herein, we describe a biobarcode and an enzyme-assisted DNA recycling hybrid amplification strategy for naked eye detection of sub-picomolar carcinoembryonic antigen (CEA), a cancer biomarker. The presence of CEA and the corresponding antibodies results in the formation of immunocomplexes and the capture of the biobarcodes in a microplate. The massive barcode DNAs released from the biobarcodes hybridize with the G-quadruplex inactive hairpin DNA probes and form catalytic nicking sites for N.BstNBI endonuclease , which cleaves the barcode DNA/hairpin partial dsDNA, releases the G-quadruplex active sequences and recycles the barcode DNA. Due to the barcode DNA recycling process, numerous G-quadruplex active sequences are generated and associate with hemin to form peroxidase mimicking enzymes, which convert colorless ABTS 2− to green color intensified ABTS − to achieve naked eye detection of CEA down to 0.025 ng mL −1 (0.14 pM). The naked eye detection strategy reported herein can be applied also to complicated serum sample matrix , making this approach hold great promise for point-of-care diagnostic applications.
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BIOSENSORS & BIOELECTRONICS
·
2013
·
IF 5.437
Bingqian Liu,Bing Zhang,Guonan Chen,Huanghao Yang,Dianping Tang
A new flow-through electrochemical aptasensor was designed for ultrasensitive monitoring of adenosine triphosphate (ATP) by coupling microvalve-programmable capillary column with CdS-functionalized DNA concatamer for signal amplification . Initially, a layer of primary DNA-conjugated polyacrylamide hydrogel was covalently linked onto the internal surface of capillary column, and then an automated sequenctial injection format with a syringe pump was employed for development of the programmable capillary-based aptasensor. In the presence of target DNA aptamer, the immobilized primary DNA hybridized with partial bases of the aptamer. The excess aptamer fregment could trigger the formation of DNA concatamer between auxiliary DNA 1 and CdS-labeled auxiliary DNA 2 . Upon target ATP introduction, a specific ATP-aptamer reaction was excuated, thereby resulting in the release of CdS-functionalized DNA concatamer from the capillary. Subsenquent anodic stripping voltammetric detection of cadmium released under acidic conditions from the released CdS nanoparticles could be conducted in a homemade detection cell. Under optimal conditions, the dynamic concentration range spanned from 0.1 pM to 10 nM ATP with a detection limit of 0.06 pM ATP. The electrochemical aptasensor showed good reproducibility, selectivity, and stability. In addition, the methodology was evaluated for the analysis of ATP spiked serum samples, and the recoveries was 81–140%.
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Nanoscale
·
2013
·Vol. 5
·期 22
·10982-10988
·
IF 6.233
Lijian Chen,Kaifang Sun,Peipei Li,Xianzhong Fan,Jianchao Sun,Shiyun Ai
LDH nanosheets were obtained via continuous impaction and exfoliation by herring sperm DNA molecules using a constant vibration method. DNA–LDH nanohybrids were composed by electrostatic forces and they exhibited DNA-enhanced peroxidase-like activity. The morphology and structure of DNA–LDH nanohybrids were analyzed by transmission electron microscopy (TEM), selected-area electron diffraction (SAED), X-ray diffraction (XRD), and atomic force microscopy (AFM) characterization. On the basis of the high catalytic activity of DNA/CuAl–LDH nanosheets, a rapid, sensitive, and convenient approach was developed for colorimetric detection of H2O2 and blood glucose. This method can be potentially applied in medical diagnostics and biotechnology fields.
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ACS Applied Materials & Interfaces
·
2015
·
IF 6.723
Cuiyun Yang,Kai Shi,Baoting Dou,Yun Xiang,Yaqin Chai,Ruo Yuan
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 electrochemical detection of microRNA (miRNA-199a) is described. The target miRNA-199a hybridizes with the partial dsDNA probes to initiate the target-assisted polymerization nicking reaction (TAPNR) amplification to produce massive intermediate sequences, which can be captured on the sensing electrode by the self-assembled DNA secondary probes. 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. DNA-templated synthesis of AgNCs can be realized by subsequent incubation of the dsDNA polymer-modified electrode with AgNO3 and sodium borohydride. With this integrated TAPNR and HCR dual amplification strategy, the amount of in situ synthesized AgNCs is dramatically enhanced, leading to substantially amplified current response for highly sensitive detection of miRNA-199a down to 0.64 fM. In addition, the developed method also shows high selectivity toward the target miRNA-199a. 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.
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ANALYTICAL CHEMISTRY
·
2015
·
IF 5.636
Feifei Xu,Ting Yang,Yun Chen
The distorted and unique expression of microRNAs (miRNAs) in cancer makes them an attractive source of biomarkers. However, one of prerequisites for the application of miRNAs in clinical practice is to accurately profile their expression. Currently available assays normally require pre-enrichment, amplification, and labeling steps, and most of them are semiquantitative. In this study, we converted the signal of target miR-21 into reporter peptide by a DNA-peptide probe and the reporter peptide was ultimately quantified using LC-MS/MS-based targeted proteomics. Specifically, substrate peptide GDKAVLGVDPFR containing reporter peptide AVLGVDPFR and tryptic cleavage site (lysine at position 3) was first designed, followed by the conjugation with DNA sequence that was complementary to miR-21. The newly formed DNA-peptide probe was then hybridized with miR-21, which was biotinylated and attached to streptavidin agarose in advance. After trypsin digestion, the reporter peptide was released and monitored by a targeted proteomics assay. The obtained limit of quantification (LOQ) was 1 pM, and the detection dynamic range spanned ∼5 orders of magnitude. Using this assay, the developed quasi-targeted proteomics approach was applied to determine miR-21 level in breast cells and tissue samples. Finally, qRT-PCR was also performed for a comparison. This report grafted the strategy of targeted proteomics into miRNA quantification.
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ACS Chemical Biology
·
2017
·
IF 4.995
Jun Xiong,Xiaona Liu,Qing-Yun Cheng,Shan Xiao,Lai-Xin Xia,Bi-Feng Yuan,Yu-Qi Feng
Toxic heavy metals have been considered to be harmful environmental contaminations. The molecular mechanisms of heavy-metals-induced cytotoxicity and carcinogenicity are still not well elucidated. Previous reports showed exposures to toxic heavy metals can cause a change of DNA cytosine methylation (5-methylcytosine, 5-mC). However, it is still not clear whether heavy metals have effects on the recently identified new epigenetic marks in both DNA and RNA, i.e., 5-hydroxymethylcytosine (5-hmC), 5-formylcytosine (5-foC), and 5-carboxylcytosine (5-caC). Here, we established a chemical labeling strategy in combination with liquid chromatography–electrospray ionization–mass spectrometry (LC-ESI-MS/MS) analysis for highly sensitive detection of eight modified cytidines in DNA and RNA. The developed method allowed simultaneous detection of all eight modified cytidines with improved detection sensitivities of 128–443-fold. Using this method, we demonstrated that the levels of 5-hmC, 5-foC, and 5-caC significantly decreased in both the DNA and RNA of mouse embryonic stem (ES) cells while exposed to arsenic (As), cadmium (Cd), chromium (Cr), and antimony (Sb). In addition, we found that treatments by heavy metals induced a decrease of the activities of 10–11 translocation (Tet) proteins. Furthermore, we revealed that a content change of metabolites occurring in the tricarboxylic acid cycle may be responsible for the decline of the derivatives of 5-mC. Our study shed light on the epigenetic effects of heavy metals, especially for the induced decline of the derivatives of 5-mC in both DNA and RNA.
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