DNA Hybridization Detection Based on Resonance Frequency Readout in Graphene on Au SPR Biosensor

This paper demonstrates a numerical modeling of surface plasmon resonance (SPR) biosensor for detecting DNA hybridization by recording the resonance frequency characteristics (RFC). The proposed sensor is designed based on graphene material as biomolecular recognition elements (BRE) and the sharp SPR curve of gold (Au). Numerical analysis shows that the variation of RFC for mismatched DNA strands is quiet negligible whereas that for complementary DNA strands is considerably countable. Here, graphene is used to perform faster immobilization between target DNA and probe DNA. The usage of graphene also changes the RFC that ensure hybridization of DNA event by utilizing its optochemical property. In addition, proposed sensor successfully distinguishes between hybridization and single-nucleotide polymorphisms (SNP) by observing the variation level of RFC and maximum transmittance. Therefore, the proposed frequency readout based SPR sensor could potentially open a new window of detection for biomolecular interactions. We also highlight the advantage of using graphene sublayer by performing the sensitivity analysis. Sandwiching of each graphene sublayer enhances 95% sensitivity comparing with conventional SPR sensor.

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FDTD Analysis Fiber Optic SPR Biosensor for DNA Hybridization: A Numerical Demonstration with Graphene

Journal of Materials and Applications ◽

10.32732/jma.2019.8.1.13 ◽

2019 ◽

Vol 8 (1) ◽

pp. 13-19 ◽

Cited By ~ 1

Author(s):

Md. Muztahidul Islam ◽

Md. Mohaiminul Islam ◽

Youshuf C. Shimul ◽

Azizur Rahman ◽

A. Akib Ruhe ◽

...

Keyword(s):

Dna Hybridization ◽

Graphene Layer ◽

Fiber Optic ◽

Biomedical Application ◽

Fdtd Method ◽

Attenuated Total Reflection ◽

Mismatched Dna ◽

Gold Thin Film ◽

Spr Biosensor ◽

Dna Strands

This article illustrates a design and finite difference time domain (FDTD) method based on analysis of fiber optic surface plasmon resonance (SPR) biosensor for biomedical application especially for DNA-DNA hybridization. The fiber cladding at the middle portion is constructed with the proposed hybrid of gold (Au), graphene, and a sensing medium. This sensor can be recognized adsorption of DNA biomolecules onto sensing medium of PBS saline using attenuated total reflection (ATR) technique. The refractive index (RI) is varied owing to the adsorption of different concentration of biomolecules. Result states that the sensitivity with a monolayer of graphene will be improved up to 40% than bare graphene layer. Owing to increased adsorption capability of DNA molecules on graphene, sensitivity increases compared to the conventional gold thin film SPR biosensor. Numerical analysis shows that the variation of the SPR angle for mismatched DNA strands is quite negligible, whereas that for complementary DNA strands is considerable, which is essential for proper detection of DNA hybridization. Finally, the effect of Electric field distribution on inserting graphene layer is analyzed incorporating the FDTD technique by using Lumerical FDTD solution software.

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BEAR reveals that increased fidelity variants can successfully reduce the mismatch tolerance of adenine but not cytosine base editors

Nature Communications ◽

10.1038/s41467-021-26461-y ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

András Tálas ◽

Dorottya A. Simon ◽

Péter I. Kulcsár ◽

Éva Varga ◽

Sarah L. Krausz ◽

...

Keyword(s):

High Throughput ◽

Genome Engineering ◽

Sequence Context ◽

Base Editing ◽

Target Dna ◽

Dna Strands ◽

Deaminase Domain ◽

Target Effects ◽

Target Sets

AbstractAdenine and cytosine base editors (ABE, CBE) allow for precision genome engineering. Here, Base Editor Activity Reporter (BEAR), a plasmid-based fluorescent tool is introduced, which can be applied to report on ABE and CBE editing in a virtually unrestricted sequence context or to label base edited cells for enrichment. Using BEAR-enrichment, we increase the yield of base editing performed by nuclease inactive base editors to the level of the nickase versions while maintaining significantly lower indel background. Furthermore, by exploiting the semi-high-throughput potential of BEAR, we examine whether increased fidelity SpCas9 variants can be used to decrease SpCas9-dependent off-target effects of ABE and CBE. Comparing them on the same target sets reveals that CBE remains active on sequences, where increased fidelity mutations and/or mismatches decrease the activity of ABE. Our results suggest that the deaminase domain of ABE is less effective to act on rather transiently separated target DNA strands, than that of CBE explaining its lower mismatch tolerance.

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Nanobody Detection of Standard Fluorescent Proteins Enables Multi-Target DNA-PAINT with High Resolution and Minimal Displacement Errors

10.1101/500298 ◽

2018 ◽

Cited By ~ 1

Author(s):

Shama Sograte-Idrissi ◽

Nazar Oleksiievets ◽

Sebastian Isbaner ◽

Mariana Eggert-Martinez ◽

Jörg Enderlein ◽

...

Keyword(s):

Fluorescent Proteins ◽

Super Resolution ◽

Resolving Power ◽

Acquisition Time ◽

Efficient Manner ◽

Target Dna ◽

Multiplexed Imaging ◽

Dna Strands ◽

Linkage Error ◽

20 Nm

AbstractDNA-PAINT is a rapidly developing fluorescence super-resolution technique which allows for reaching spatial resolutions below 10 nm. It also enables the imaging of multiple targets in the same sample. However, using DNA-PAINT to observe cellular structures at such resolution remains challenging. Antibodies, which are commonly used for this purpose, lead to a displacement between the target protein and the reporting fluorophore of 20-25 nm, thus limiting the resolving power. Here, we used nanobodies to minimize this linkage error to ~4 nm. We demonstrate multiplexed imaging by using 3 nanobodies, each able to bind to a different family of fluorescent proteins. We couple the nanobodies with single DNA strands via a straight forward and stoichiometric chemical conjugation. Additionally, we built a versatile computer-controlled microfluidic setup to enable multiplexed DNA-PAINT in an efficient manner. As a proof of principle, we labeled and imaged proteins on mitochondria, the Golgi apparatus, and chromatin. We obtained super-resolved images of the 3 targets with 20 nm resolution, and within only 35 minutes acquisition time.

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Sequence-Specific Detection of DNA Strands Using a Solid-State Nanopore Assisted by Microbeads

Micromachines ◽

10.3390/mi11121097 ◽

2020 ◽

Vol 11 (12) ◽

pp. 1097

Author(s):

Yin Zhang ◽

Zengdao Gu ◽

Jiabin Zhao ◽

Liying Shao ◽

Yajing Kan

Keyword(s):

Low Cost ◽

Local Concentration ◽

Dna Molecules ◽

Dna Translocation ◽

Specific Sequence ◽

Free Dna ◽

Target Dna ◽

Dna Strands ◽

Ionic Pulse ◽

Biotinylated Probes

Simple, rapid, and low-cost detection of DNA with specific sequence is crucial for molecular diagnosis and therapy applications. In this research, the target DNA molecules are bonded to the streptavidin-coated microbeads, after hybridizing with biotinylated probes. A nanopore with a diameter significantly smaller than the microbeads is used to detect DNA molecules through the ionic pulse signals. Because the DNA molecules attached on the microbead should dissociate from the beads before completely passing through the pore, the signal duration time for the target DNA is two orders of magnitude longer than free DNA. Moreover, the high local concentration of target DNA molecules on the surface of microbeads leads to multiple DNA molecules translocating through the pore simultaneously, which generates pulse signals with amplitude much larger than single free DNA translocation events. Therefore, the DNA molecules with specific sequence can be easily identified by a nanopore sensor assisted by microbeads according to the ionic pulse signals.

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Rapid DNA hybridization based on ac field focusing of magnetically labeled target DNA

Applied Physics Letters ◽

10.1063/1.1984090 ◽

2005 ◽

Vol 87 (1) ◽

pp. 013901 ◽

Cited By ~ 34

Author(s):

H. A. Ferreira ◽

N. Feliciano ◽

D. L. Graham ◽

L. A. Clarke ◽

M. D. Amaral ◽

...

Keyword(s):

Dna Hybridization ◽

Target Dna ◽

Ac Field

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A turn-on dual emissive nucleobase sensitive to mismatches and duplex conformational changes

RSC Advances ◽

10.1039/c6ra19061h ◽

2016 ◽

Vol 6 (90) ◽

pp. 87142-87146 ◽

Cited By ~ 4

Author(s):

Krishna Gavvala ◽

Nicolas P. F. Barthes ◽

Dominique Bonhomme ◽

Anne Sophie Dabert-Gay ◽

Delphine Debayle ◽

...

Keyword(s):

Single Nucleotide Polymorphisms ◽

Conformational Changes ◽

Dna Hybridization ◽

Nucleotide Polymorphisms ◽

Single Nucleotide ◽

Turn On ◽

Fluorescent Nucleoside

Herein, we demonstrate the on–off dual emissive behaviour of a fluorescent nucleoside sensitive towards DNA hybridization and conformational changes as well as detection of single nucleotide polymorphisms.

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A Novel Inductive Coupled Plasma Mass Spectrometry Gene Detection Method Based on AuNPs and Bio-Barcode Signal Amplification

Nanoscience and Nanotechnology Letters ◽

10.1166/nnl.2019.2933 ◽

2019 ◽

Vol 11 (5) ◽

pp. 638-644 ◽

Cited By ~ 3

Author(s):

Pengfei Jiang ◽

Chomba Haji ◽

Xiufang Ye ◽

Menglei Chang ◽

Wenjun Li ◽

...

Keyword(s):

Mass Spectrometry ◽

Signal Amplification ◽

Limit Of Detection ◽

High Sensitivity ◽

Mismatched Dna ◽

Icp Ms ◽

Target Dna ◽

Cancer Related Gene ◽

Plasma Mass Spectrometry ◽

Double Base

An inductive coupled plasma mass spectrometry (ICP-MS) method based on gold nanoparticles (AuNPs) and bio-barcode signal amplification was presented for the sensitive detection of gastric cancer related gene. The target DNA, magnetic nanoprobes and SiO2/AuNPs barcode probes were hybridized to form a "sandwich" structure, then a large amount of AuNPs which were amplified by bio-barcode technology were detected by ICP-MS. This method presented a limit of detection as low as 1 fM. The ratio of the background-subtracted 197 Au signals for totally complementary DNA, single-base mismatched DNA, double-base mismatched DNA and totally mismatched DNA was 115:30:16:1, respectively. It's suggested that the complementary and mismatched DNA can be distinguished clearly. This novel ICP-MS biosensor is prospective in DNA detection with high sensitivity and specificity.

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Label-Free Dengue Detection Utilizing PNA/DNA Hybridization Based on the Aggregation Process of Unmodified Gold Nanoparticles

Journal of Nanomaterials ◽

10.1155/2014/839286 ◽

2014 ◽

Vol 2014 ◽

pp. 1-5 ◽

Cited By ~ 5

Author(s):

Samsulida Abdul Rahman ◽

Rafidah Saadun ◽

Nur Ellina Azmi ◽

Nurhayati Ariffin ◽

Jaafar Abdullah ◽

...

Keyword(s):

Gold Nanoparticles ◽

Dna Hybridization ◽

Peptide Nucleic Acid ◽

Virus Detection ◽

Label Free ◽

Color Changes ◽

Phosphate Backbone ◽

Single Base Mismatch ◽

Target Dna ◽

Base Mismatch

A label-free optical detection method based on PNA/DNA hybridization using unmodified gold nanoparticles (AuNPs) for dengue virus detection has been successfully developed. In this study, no immobilization method is involved and the hybridization of PNA/DNA occurs directly in solution. Unmodified AuNPs undergo immediate aggregation in the presence of neutral charge peptide nucleic acid (PNA) due to the coating of PNA on AuNPs surface. However, in the presence of complementary targets DNA, the hybridization of PNA probe with target DNA forms negatively charged complexes due to the negatively charged phosphate backbone of the target DNA. The negatively charged complexes adsorbed onto the AuNPs surface ensure sufficient charge repulsion, need for AuNPs dispersion, and stability in solution. The detection procedure is a naked eye method based on immediate color changes and also through UV-vis adsorption spectra. The selectivity of the proposed method was studied successfully by single base mismatch and noncomplementary target DNA.

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DNA Ligase-Based Strategy for Quantifying Heterogeneous DNA Methylation without Sequencing

Clinical Chemistry ◽

10.1373/clinchem.2014.227546 ◽

2015 ◽

Vol 61 (1) ◽

pp. 163-171 ◽

Cited By ~ 17

Author(s):

Eugene J H Wee ◽

Sakandar Rauf ◽

Muhammad J A Shiddiky ◽

Alexander Dobrovic ◽

Matt Trau

Keyword(s):

Dna Methylation ◽

Single Molecule ◽

Region Of Interest ◽

Suppressor Gene ◽

Fluorescent Detection ◽

Routine Practice ◽

Dna Ligases ◽

Mismatched Dna ◽

Dna Strands ◽

Analytical Accuracy

Abstract BACKGROUND DNA methylation is a potential source of disease biomarkers. Typically, methylation levels are measured at individual cytosine/guanine (CpG) sites or over a short region of interest. However, regions of interest often show heterogeneous methylation comprising multiple patterns of methylation (epialleles) on individual DNA strands. Heterogeneous methylation is largely ignored because digital methods are required to deconvolute these usually complex patterns of epialleles. Currently, only single-molecule approaches, such as next generation sequencing (NGS), can provide detailed epiallele information. Because NGS is not yet feasible for routine practice, we developed a single-molecule–like approach, named for epiallele quantification (EpiQ). METHODS EpiQ uses DNA ligases and the enhanced thermal instability of short (≤19 bases) mismatched DNA probes for the relative quantification of epialleles. The assay was developed using fluorescent detection on a gel and then adapted for electrochemical detection on a microfabricated device. NGS was used to validate the analytical accuracy of EpiQ. RESULTS In this proof of principle study, EpiQ detected with 90%–95% specificity each of the 8 possible epialleles for a 3-CpG cluster at the promoter region of the CDKN2B (p15) tumor suppressor gene. EpiQ successfully profiled heterogeneous methylation patterns in clinically derived samples, and the results were cross-validated with NGS. CONCLUSIONS EpiQ is a potential alternative tool for characterizing heterogeneous methylation, thus facilitating its use as a biomarker. EpiQ was developed on a gel-based assay but can also easily be adapted for miniaturized chip-based platforms.

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Mechanistic Insights into theCis-andTrans-acting Deoxyribonuclease Activities of Cas12a

10.1101/353748 ◽

2018 ◽

Cited By ~ 2

Author(s):

Daan C. Swarts ◽

Martin Jinek

Keyword(s):

Genome Editing ◽

Dna Cleavage ◽

Cleavage Product ◽

List Type ◽

Francisella Novicida ◽

Ssdna Binding ◽

Double Stranded Dna ◽

Target Dna ◽

Dna Strands ◽

Dna Strand

HIGHLIGHTSTarget ssDNA binding allosterically induces unblocking of the RuvC active sitePAM binding facilitates unwinding of dsDNA targetsNon-target DNA strand cleavage is prerequisite for target DNA strand cleavageAfter DNA cleavage, Cas12a releases the PAM-distal DNA productSUMMARYCRISPR-Cas12a (Cpf1) is an RNA-guided DNA-cutting nuclease that has been repurposed for genome editing. Upon target DNA binding, Cas12a cleaves both the target DNA incisand non-target single stranded DNAs (ssDNA) intrans.To elucidate the molecular basis for both deoxyribonuclease cleavage modes, we performed structural and biochemical studies onFrancisella novicidaCas12a. We show how crRNA-target DNA strand hybridization conformationally activates Cas12a, triggering itstrans-acting, non-specific, single-stranded deoxyribonuclease activity. In turn,cis-cleavage of double-stranded DNA targets is a result of PAM-dependent DNA duplex unwinding and ordered sequential cleavage of the non-target and target DNA strands. Cas12a releases the PAM-distal DNA cleavage product and remains bound to the PAM-proximal DNA cleavage product in a catalytically competent,trans-active state. Together, these results provide a revised model for the molecular mechanism of Cas12a enzymes that explains theircis- andtrans-acting deoxyribonuclease activities, and additionally contribute to improving Cas12a-based genome editing.

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