Quencher-free molecular beacon tethering 7-hydroxycoumarin detects targets through protonation/deprotonation

Kashida, Hiromu; Yamaguchi, Kyohei; Hara, Yuichi; Asanuma, Hiroyuki

doi:10.1016/j.bmc.2012.05.052

Cited by 16 publications

(5 citation statements)

References 35 publications

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“…Variations of such MB probes are shown in Figure 10. For example, Kashida et al described quencher-free MB probe containing 7-hydroxycoumarin as a fluorescent reporter [113] (Figure 10(a)). Interestingly, the pKa of in-stem bound dye was 1.2 units higher than that in a single-stranded DNA.…”

Section: Challengesmentioning

confidence: 99%

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An Elegant Biosensor Molecular Beacon Probe: Challenges and Recent Solutions

Kolpashchikov

2012

Scientifica

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Molecular beacon (MB) probes are fluorophore- and quencher-labeled short synthetic DNAs folded in a stem-loop shape. Since the first report by Tyagi and Kramer, it has become a widely accepted tool for nucleic acid analysis and triggered a cascade of related developments in the field of molecular sensing. The unprecedented success of MB probes stems from their ability to detect specific DNA or RNA sequences immediately after hybridization with no need to wash out the unbound probe (instantaneous format). Importantly, the hairpin structure of the probe is responsible for both the low fluorescent background and improved selectivity. Furthermore, the signal is generated in a reversible manner; thus, if the analyte is removed, the signal is reduced to the background. This paper highlights the advantages of MB probes and discusses the approaches that address the challenges in MB probe design. Variations of MB-based assays tackle the problem of stem invasion, improve SNP genotyping and signal-to-noise ratio, as well as address the challenges of detecting folded RNA and DNA.

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Section: Challengesmentioning

confidence: 99%

“…(a) Quencher-free in-stem MB probe developed by Kashida et al [113]. (b) “Smart” probes take advantage of multiple guanine residues located in the stem to quench the fluorescence of a fluorophore dye [114].…”

Section: Figurementioning

confidence: 99%

An Elegant Biosensor Molecular Beacon Probe: Challenges and Recent Solutions

Kolpashchikov

2012

Scientifica

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“…The Asanuma group designed a quencher-free MB by introducing 7-hydroxycoumarin ( X ) into the stem region ( Figure 17 a) [ 87 ]. X showed quenched fluorescence upon protonation.…”

Section: Microenvironment-sensitive Probesmentioning

confidence: 99%

Single-Labeled Oligonucleotides Showing Fluorescence Changes upon Hybridization with Target Nucleic Acids

Hwang

2018

Molecules

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Sequence-specific detection of nucleic acids has been intensively studied in the field of molecular diagnostics. In particular, the detection and analysis of single-nucleotide polymorphisms (SNPs) is crucial for the identification of disease-causing genes and diagnosis of diseases. Sequence-specific hybridization probes, such as molecular beacons bearing the fluorophore and quencher at both ends of the stem, have been developed to enable DNA mutation detection. Interestingly, DNA mutations can be detected using fluorescently labeled oligonucleotide probes with only one fluorophore. This review summarizes recent research on single-labeled oligonucleotide probes that exhibit fluorescence changes after encountering target nucleic acids, such as guanine-quenching probes, cyanine-containing probes, probes containing a fluorophore-labeled base, and microenvironment-sensitive probes.

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“…Use of pH-dependent probes has been already described for selective imaging of cancer cells with fluorescent cancer cell-targeted antibodies [207]. In the case of molecular beacon, a pH-sensitive fluorophore 7-hydroxycoumarin was inserted as a nucleobase analogue into the hairpin stem [208]. The pKa of 7-hydroxycoumarin depends on the microenvironment: in a single strand it has pKa 8.8, whereas being intercalated it changes its pKa to 10.…”

Section: Probes For Rna and Single-stranded Dna Imaging By Fluoresmentioning

confidence: 99%

Fluorescent Probes for Nucleic Acid Visualization in Fixed and Live Cells

et al. 2013

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This review analyses the literature concerning non-fluorescent and fluorescent probes for nucleic acid imaging in fixed and living cells from the point of view of their suitability for imaging intracellular native RNA and DNA. Attention is mainly paid to fluorescent probes for fluorescence microscopy imaging. Requirements for the target-binding part and the fluorophore making up the probe are formulated. In the case of native double-stranded DNA, structure-specific and sequence-specific probes are discussed. Among the latest, three classes of dsDNA-targeting molecules are described: (i) sequence-specific peptides and proteins; (ii) triplex-forming oligonucleotides and (iii) polyamide oligo(N-methylpyrrole/N-methylimidazole) minor groove binders. Polyamides seem to be the most promising targeting agents for fluorescent probe design, however, some technical problems remain to be solved, such as the relatively low sequence specificity and the high background fluorescence inside the cells. Several examples of fluorescent probe applications for DNA imaging in fixed and living cells are cited. In the case of intracellular RNA, only modified oligonucleotides can provide such sequence-specific imaging. Several approaches for designing fluorescent probes are considered: linear fluorescent probes based on modified oligonucleotide analogs, OPEN ACCESSMolecules 2013, 18 15358 molecular beacons, binary fluorescent probes and template-directed reactions with fluorescence probe formation, FRET donor-acceptor pairs, pyrene excimers, aptamers and others. The suitability of all these methods for living cell applications is discussed.

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Quencher-free molecular beacon tethering 7-hydroxycoumarin detects targets through protonation/deprotonation

Cited by 16 publications

References 35 publications

An Elegant Biosensor Molecular Beacon Probe: Challenges and Recent Solutions

An Elegant Biosensor Molecular Beacon Probe: Challenges and Recent Solutions

Single-Labeled Oligonucleotides Showing Fluorescence Changes upon Hybridization with Target Nucleic Acids

Fluorescent Probes for Nucleic Acid Visualization in Fixed and Live Cells

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