-
Nanoscale Sep 2023Identification of non-amplified DNA sequences and single-base mutations is essential for molecular biology and genetic diagnostics. This paper reports a novel sensor...
Identification of non-amplified DNA sequences and single-base mutations is essential for molecular biology and genetic diagnostics. This paper reports a novel sensor consisting of electrochemically-gated graphene coplanar waveguides coupled with a microfluidic channel. Upon exposure to analytes, propagation of electromagnetic waves in the waveguides is modified as a result of interactions with the fringing field and modulation of graphene dynamic conductivity resulting from electrostatic gating. Probe DNA sequences are immobilised on the graphene surface, and the sensor is exposed to DNA sequences which either perfectly match the probe, contain a single-base mismatch or are unrelated. By monitoring the scattering parameters at frequencies between 50 MHz and 50 GHz, unambiguous and reproducible discrimination of the different strands is achieved at concentrations as low as one attomole per litre (1 aM). By controlling and synchronising frequency sweeps, electrochemical gating, and liquid flow in the microfluidic channel, the sensor generates multidimensional datasets. Advanced data analysis techniques are utilised to take full advantage of the richness of the dataset. A classification accuracy >97% between all three sequences is achieved using different Machine Learning models, even in the presence of simulated noise and low signal-to-noise ratios. The sensor exceeds state-of-the-art sensitivity of field-effect transistors and microwave sensors for the identification of single-base mismatches.
Topics: Graphite; Microwaves; Biosensing Techniques; DNA Probes; Signal-To-Noise Ratio
PubMed: 37682040
DOI: 10.1039/d3nr01239e -
Analytical Chemistry Dec 2022Traditional spherical nucleic acids (SNAs) based on gold nanoparticles (AuNPs) assembled through Au-S covalent bonds are widely used in DNA-programmable assembly,...
Traditional spherical nucleic acids (SNAs) based on gold nanoparticles (AuNPs) assembled through Au-S covalent bonds are widely used in DNA-programmable assembly, biosensing, imaging, and therapeutics. However, biological thiols and other chemical substances can break the Au-S bonds and cause response distortion during the application process, specifically in cell environments. Herein, we report a new type of SNAs based on 2'-fluorinated DNA-functionalized AuNPs with excellent colloidal stability under high salt conditions (up to 1 M NaCl) and over a broad pH range (1-14), as well as resistance to biothiols. The fluorinated spherical nucleic acid probe (Au/FDNA probe) could detect targeted cancer cells with high fidelity. Compared to the traditional thiolated DNA-functionalized AuNP probe (Au-SDNA probe), the Au/FDNA probe exhibited a higher sensitivity to the target and a lower signal-to-background ratio. Furthermore, the Au/FDNA probe could discriminate target cancer cells in a mixed culture system. Using the proposed FDNA functionalization method, previously developed SNAs based on AuNPs could be directly adapted, which might open a new avenue for the design and application of SNAs.
Topics: Gold; Metal Nanoparticles; DNA; Nucleic Acids; DNA Probes; Biosensing Techniques
PubMed: 36519891
DOI: 10.1021/acs.analchem.2c04294 -
International Journal of Molecular... Jul 2021Oligonucleotides fluorescence in situ hybridization (Oligo-FISH) is an emerging technology and is an important tool in research areas such as detection of chromosome... (Review)
Review
Oligonucleotides fluorescence in situ hybridization (Oligo-FISH) is an emerging technology and is an important tool in research areas such as detection of chromosome variation, identification of allopolyploid, and deciphering of three-dimensional (3D) genome structures. Based on the demand for highly efficient oligo probes for oligo-FISH experiments, increasing numbers of tools have been developed for probe design in recent years. Obsolete oligonucleotide design tools have been adapted for oligo-FISH probe design because of their similar considerations. With the development of DNA sequencing and large-scale synthesis, novel tools have been designed to increase the specificity of designed oligo probes and enable genome-scale oligo probe design, which has greatly improved the application of single copy oligo-FISH. Despite this, few studies have introduced the development of the oligo-FISH probe design tools and their application in FISH experiments systematically. Besides, a comprehensive comparison and evaluation is lacking for the available tools. In this review, we provide an overview of the oligo-FISH probe design process, summarize the development and application of the available tools, evaluate several state-of-art tools, and eventually provide guidance for single copy oligo-FISH probe design.
Topics: Chromosomes, Plant; DNA Probes; Fluorescence; Genome; In Situ Hybridization, Fluorescence; Oligonucleotide Probes; Oligonucleotides; Repetitive Sequences, Nucleic Acid; Research Design; Sequence Analysis, DNA
PubMed: 34281175
DOI: 10.3390/ijms22137124 -
Analytical and Bioanalytical Chemistry Mar 2016The development of rapid, cost-effective DNA detection methods for molecular diagnostics at the point-of-care (POC) has been receiving increasing interest. This article... (Review)
Review
The development of rapid, cost-effective DNA detection methods for molecular diagnostics at the point-of-care (POC) has been receiving increasing interest. This article reviews several DNA detection techniques based on plasmonic-active nanochip platforms developed in our laboratory over the last 5 years, including the molecular sentinel-on-chip (MSC), the multiplex MSC, and the inverse molecular sentinel-on-chip (iMS-on-Chip). DNA probes were used as the recognition elements, and surface-enhanced Raman scattering (SERS) was used as the signal detection method. Sensing mechanisms were based on hybridization of target sequences and DNA probes, resulting in a distance change between SERS reporters and the nanochip's plasmonic-active surface. As the field intensity of the surface plasmon decays exponentially as a function of distance, the distance change in turn affects SERS signal intensity, thus indicating the presence and capture of the target sequences. Our techniques were single-step DNA detection techniques. Target sequences were detected by simple delivery of sample solutions onto DNA probe-functionalized nanochips and measuring the SERS signal after appropriate incubation times. Target sequence labeling or washing to remove unreacted components was not required, making the techniques simple, easy-to-use, and cost-effective. The usefulness of the nanochip platform-based techniques for medical diagnostics was illustrated by the detection of host genetic biomarkers for respiratory viral infection and of the dengue virus gene.
Topics: Animals; Biosensing Techniques; DNA; DNA Probes; Equipment Design; Humans; Oligonucleotide Array Sequence Analysis; Spectrum Analysis, Raman
PubMed: 26547189
DOI: 10.1007/s00216-015-9121-4 -
Angewandte Chemie (International Ed. in... Feb 2022The cell membrane is a dynamic and heterogeneous structure composed of distinct sub-compartments. Within these compartments, preferential interactions occur among...
The cell membrane is a dynamic and heterogeneous structure composed of distinct sub-compartments. Within these compartments, preferential interactions occur among various lipids and proteins. Currently, it is still challenging to image these short-lived membrane complexes, especially in living cells. In this work, we present a DNA-based probe, termed "DNA Zipper", which allows the membrane order and pattern of transient interactions to be imaged in living cells using standard fluorescence microscopes. By fine-tuning the length and binding affinity of DNA duplex, these probes can precisely extend the duration of membrane lipid interactions via dynamic DNA hybridization. The correlation between membrane order and the activation of T-cell receptor signaling has also been studied. These programmable DNA probes function after a brief cell incubation, which can be easily adapted to study lipid interactions and membrane order during different membrane signaling events.
Topics: Animals; Cell Membrane; DNA Probes; Dogs; Fluorescence Resonance Energy Transfer; Fluorescent Dyes; Madin Darby Canine Kidney Cells
PubMed: 34767659
DOI: 10.1002/anie.202112033 -
Biomarkers in Medicine Jan 2023DNA probes have been widely used as diagnostic tools for translocations. This study sought to design a screening tool using ssDNA probes and chromosome conformation...
DNA probes have been widely used as diagnostic tools for translocations. This study sought to design a screening tool using ssDNA probes and chromosome conformation capture (3C) library fragment hybridization. The authors focused on developing a probe for the juxtaposed region of and . Fragments of the gene with a thiol modification (MYC-Au NP probe) were functionalized by gold nanoparticles (Au NPs). Then probes were immobilized on a nitrocellulose surface. Hybridization between DNA probes and 3C library fragments of SKW3 cells was determined by color intensity. Optimal hybridization of the 3C library sample of the cell line to probes showed higher color intensity than human umbilical vein endothelial cells. Combining 3C-based techniques and DNA-DNA hybridization can identify rearrangements in cancer cells.
Topics: Humans; Translocation, Genetic; Gold; Endothelial Cells; Metal Nanoparticles; Chromosomes; DNA Probes; DNA; Biosensing Techniques
PubMed: 37013851
DOI: 10.2217/bmm-2022-0115 -
Biosensors & Bioelectronics Jun 2024Mechanical forces play an important role in cellular communication and signaling. We developed in this study novel electrochemical DNA-based force sensors for measuring...
Mechanical forces play an important role in cellular communication and signaling. We developed in this study novel electrochemical DNA-based force sensors for measuring cell-generated adhesion forces. Two types of DNA probes, i.e., tension gauge tether and DNA hairpin, were constructed on the surface of a smartphone-based electrochemical device to detect piconewton-scale cellular forces at tunable levels. Upon experiencing cellular tension, the unfolding of DNA probes induces the separation of redox reporters from the surface of the electrode, which results in detectable electrochemical signals. Using integrin-mediated cell adhesion as an example, our results indicated that these electrochemical sensors can be used for highly sensitive, robust, simple, and portable measurements of cell-generated forces.
Topics: Biosensing Techniques; DNA; Cell Adhesion; DNA Probes; Integrins
PubMed: 38457863
DOI: 10.1016/j.bios.2024.116185 -
The Analyst Nov 2022A simple, reliable, and cost-effective method for nucleic acid detection is of increasing interest in clinical diagnostics of infectious and genetic diseases. Currently,...
A simple, reliable, and cost-effective method for nucleic acid detection is of increasing interest in clinical diagnostics of infectious and genetic diseases. Currently, enzyme-mediated methods such as polymerase chain reactions and loop-mediated isothermal amplification are the most widely used methods in the qualitative and quantitative diagnosis of long nucleic acid sequences with high sensitivity. However, a high detection sensitivity for short-length sequences remains a significant challenge because it is difficult for the primers to bind to their sequences. Our previous study demonstrated a simple, enzyme-free, and sequence-specific colorimetric detection of 24-nucleotide short target DNA at room temperature using a developed assay that combines catalytic hairpin assembly (CHA) and enzyme-linked immunosorbent assay (ELISA)-mimicking methods. In this follow-up study, we aim to design and develop DNA-based signal amplifiers, or DNA dendrons, to improve the colorimetric detection of short target cDNA in the CHA-mediated ELISA-mimicking assay. DNA dendrons are highly branched conformations synthesized by the molecular self-assembly of three DNA oligomers. The assay using DNA dendrons demonstrates an enhanced detection sensitivity with the detection of approximately 50 pM of 24-nucleotide short target cDNA, which is a 16.4-fold higher detection limit compared to that obtained without DNA dendrons under the same conditions. Thus, applications of the developed DNA dendrons as an effective signal amplifier in DNA probe-based chemiluminescence assays have the potential to improve the colorimetric detection of short target cDNA with high sensitivity for the diagnosis of different diseases.
Topics: Colorimetry; DNA, Complementary; Dendrimers; Follow-Up Studies; Luminescence; DNA; Nucleic Acid Amplification Techniques; DNA Probes; Nucleotides; Limit of Detection; Biosensing Techniques
PubMed: 36239244
DOI: 10.1039/d2an01137a -
Bioelectrochemistry (Amsterdam,... Oct 2020Applications of molecular techniques to elucidate identity or function using biomarkers still remain highly empirical and biosensors are no exception. In the present...
Applications of molecular techniques to elucidate identity or function using biomarkers still remain highly empirical and biosensors are no exception. In the present study, target-specific oligonucleotide probes for E. coli K12 were designed thermodynamically and applied in an electrochemical DNA biosensor setup. Biosensor was prepared by immobilization of a stem-loop structured probe, modified with a thiol functional group at its 5' end and a biotin molecule at its 3' end, on a gold electrode through self-assembly. Mercaptopropionic acid (MPA) was used to optimize the surface probe density of the electrode. Hybridization between the immobilized probe and the target DNA was detected via the electrochemical response of streptavidin-horseradish peroxidase in the presence of the substrate. The amperometric response showed a linear relationship with the target DNA concentration, ranging from 10 and 400 nM, with a correlation coefficient of 0.989. High selectivity and good repeatability of the biosensor showed that the thermodynamic approach to oligonucleotide probe design can be used in development of electrochemical DNA biosensors.
Topics: Biosensing Techniques; Biotin; DNA Probes; Electrochemical Techniques; Horseradish Peroxidase; Limit of Detection; Propionates; Reproducibility of Results; Streptavidin; Thermodynamics
PubMed: 32442773
DOI: 10.1016/j.bioelechem.2020.107553 -
ACS Applied Bio Materials Jan 2024Cancers remain the leading cause of mortality worldwide. It is crucial to detect cancer at an early stage for improving survival rates. Biomarkers have precise...
Cancers remain the leading cause of mortality worldwide. It is crucial to detect cancer at an early stage for improving survival rates. Biomarkers have precise implications for cancer progression. Here, we built a straightforward DNA probe system that could be activated by near-infrared light to detect dual miRNAs with a high specificity. This probe is built on the basis of upconversion nanoparticles, which could emit ultraviolet light and activate DNA probes adsorbed on the outer layer. The DNA probe system is remotely controlled through manipulation of the near-infrared (NIR) light, enabling simultaneous detection of dual miRNAs. The DNA nanosystem could be effectively endocytosed by cancer cells and reflect expression levels of dual miRNAs. Overall, this study demonstrates a promising remote-controlled DNA nanoplatform for the simultaneous detection of dual miRNAs, which has tremendous potential for precise cancer diagnostics and therapies.
Topics: Humans; MicroRNAs; Ultraviolet Rays; DNA; DNA Probes; Nanoparticles; Neoplasms
PubMed: 38151236
DOI: 10.1021/acsabm.3c01079