Development of a graphene oxide-based assay for the sequence-specific detection of double-stranded DNA molecules

Graphene oxide (GO) is a promising material for the development of cost-effective detection systems. In this work, we have devised a simple and rapid GO-based method for the sequence-specific identification of DNA molecules generated by PCR amplification. The csp genes of Escherichia coli, which sha...

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Bibliographic Details
Published in:PloS one 2017-08, Vol.12 (8), p.e0183952-e0183952
Main Authors: Giuliodori, Anna Maria, Brandi, Anna, Kotla, Shivaram, Perrozzi, Francesco, Gunnella, Roberto, Ottaviano, Luca, Spurio, Roberto, Fabbretti, Attilio
Format: Article
Language:English
Subjects:
Amplification
Biology and Life Sciences
Biosensing Techniques - methods
Complementarity
Deoxyribonucleic acid
DNA
DNA - isolation & purification
E coli
Fluorescence
Fluorescent Dyes
Genes
Graphene
Graphite - chemistry
Hybridization
Methods
Nucleotide sequence
Oligonucleotides
Oxides - chemistry
Physical Sciences
Polymerase chain reaction
Polymerase Chain Reaction - methods
Primers
Proteins
Quenching
Research and Analysis Methods
Single-stranded DNA
System effectiveness
Veterinary medicine
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Summary:Graphene oxide (GO) is a promising material for the development of cost-effective detection systems. In this work, we have devised a simple and rapid GO-based method for the sequence-specific identification of DNA molecules generated by PCR amplification. The csp genes of Escherichia coli, which share a high degree of sequence identity, were selected as paradigm DNA templates. All tested csp genes were amplified with unlabelled primers, which can be rapidly removed at the end of the PCR taking advantage of the preferential binding to GO of single-stranded versus duplex DNA molecules. The amplified DNAs (targets) were heat-denatured and hybridized to a fluorescently-labelled single strand oligonucleotide (probe), which recognizes a region of the target DNAs displaying sequence variability. This interaction is extremely specific, taking place with high efficiency only when target and probe show perfect or near perfect matching. Upon GO addition, the unbound fraction of the probe was captured and its fluorescence quenched by the GO's molecular properties. On the other hand, the probe-target complexes remained in solution and emitted a fluorescent signal whose intensity was related to their degree of complementarity.
ISSN:1932-6203
1932-6203
DOI:10.1371/journal.pone.0183952