Mutation Detection in DNA Oligonucleotides Based on a Guanine Quenching Method Coupled with Enzymatic Digestion of Single-Stranded DNA

Maruyama, Tatsuo; Shinohara, Toshimitsu; Ichinose, Hirofumi; Kitaoka, Momoko; Okamura, Nobuko; Kamiya, Noriho; Goto, Masahiro

doi:10.1007/s10529-005-3681-x

Cited by 22 publications

(14 citation statements)

References 18 publications

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“…In this experiment, the fluorescence intensity decreased upon increases in the peptide concentration (Figure 6A); however, the percent change in the observed fluorescence intensity was fairly small (7%). One possible reason for this observation is quenching of the fluorescein fluorescence by the adjacent 5′-guanosine residue [63,64]. A plot of the fraction bound versus peptide concentration and hyperbolic curve fitting gave an apparent K D of 27 ± 4 μM (R 2 = 0.97) (Figure 6B ) .…”

Section: Resultsmentioning

confidence: 99%

Selection of Peptides Targeting Helix 31 of Bacterial 16S Ribosomal RNA by Screening M13 Phage-Display Libraries

et al. 2011

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Ribosomal RNA is the catalytic portion of ribosomes, and undergoes a variety of conformational changes during translation. Structural changes in ribosomal RNA can be facilitated by the presence of modified nucleotides. Helix 31 of bacterial 16S ribosomal RNA harbors two modified nucleotides, m 2 G966 and m 5 C967, that are highly conserved among bacteria, though the degree and nature of the modifications in this region are different in eukaryotes. Contacts between helix 31 and the P-site tRNA, initiation factors, and ribosomal proteins highlight the importance of this region in translation. In this work, a heptapeptide M13 phage-display library was screened for ligands that target the wild-type, naturally modified bacterial helix 31. Several peptides, including TYLPWPA, CVRPFAL, TLWDLIP, FVRPFPL, ATPLWLK, and DIRTQRE, were found to be prevalent after several rounds of screening. Several of the peptides exhibited moderate affinity (in the high nM to low µM range) to modified helix 31 in biophysical assays, including surface OPEN ACCESSMolecules 2011, 16 1212 plasmon resonance (SPR), and were also shown to bind 30S ribosomal subunits. These peptides also inhibited protein synthesis in cell-free translation assays.

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Section: Resultsmentioning

confidence: 99%

Selection of Peptides Targeting Helix 31 of Bacterial 16S Ribosomal RNA by Screening M13 Phage-Display Libraries

et al. 2011

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“…This is probably because the quenching of ROX by the guanine base present in cGene-ROX is static and originates a non-emissive exciplex, whereas the exciplex of ROX with the other bases have higher fluorescence lifetimes. [18][19][20][21][22][23][24] After the addition of Gene-MG to cGene-ROX (in 180 mm buffer at 40 8C), the fluorescence decay curves become faster ( Figure 6), with the average lifetimes decreasing from 5.1 ns (before addition) to 4.2 ns after hybridization for all acceptor concentrations. This corresponds to an energy transfer efficiency [Eq.…”

Section: Fret Detection Of Dna Hybridization In Solutionmentioning

confidence: 99%

“…We note that the decays obtained for ROX-labeled DNA strands containing guanine could only be fitted by a sum of exponential functions because guanine is a ROX fluorescence quencher. [18][19][20][21][22][23][24] Previous results shown that the polarity of the PNIPAM shell [probed by fluorescence using dT25-Rox ( Figure 1)] is practically identical to the polarity of water at temperatures below the T VPT , but decreases sharply above T VPT because the shell dehy-drates. [25] The variation in shell properties strongly influences the distribution and dynamics of DNA adsorbed onto the shell of the nanoparticles at a temperature below T VPT .…”

Section: Introductionmentioning

confidence: 99%

DNA Hybridization in Thermoresponsive Polymer Nanoparticles

2010

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We achieve very high hybridization efficiencies by using a new method to immobilize DNA strands on the surface of thermoresponsive polymer nanoparticles. Hybridization efficiencies of about 70 % are obtained between the DNA immobilized in the particles and a complementary strand in solution, even at very low ionic strengths (1 mM). The polymer nanoparticles have a glassy poly(methylmethacrylate) (PMMA) core and a thermoresponsive shell of poly(N-isopropylacrylamide) (PNIPAM) containing positive charges. After a DNA strand labeled with a fluorescence probe is loaded onto the particles at room temperature, the temperature is increased above the volume phase transition temperature of the PNIPAM shell, TVPT approximately 28 degrees C. The collapse of the particle shell immobilizes the DNA while maintaining its availability for hybridization with a complementary strand. Förster resonance energy transfer (FRET) is used to detect the hybridization with a complementary DNA strand labeled with a FRET acceptor probe.

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“…The TBA has a random-coil structure in the absence of Pb 2+ . Because the internally labeled TMR can intramolecularly interact with the adjacent guanine bases by photo-induced electron transfer (PET) mechanism due to its good electron donating property [67][68][69][70][71], the rotation of TMR is firmly restricted, which results in the aptamer probe having large FA value. In the presence of Pb 2+ , because Pb 2+ can interact with aptamer probe and induce conformational change from a random-coil structure into a highly-ordered G-quadruplex [65,72], this would weaken or eliminate the intramolecular interaction between the TMR and the adjacent guanine bases involved in formation of the G-quadruplex.…”

Section: Principle Of Sensing Pb 2+ By Famentioning

confidence: 99%

A sensitive fluorescence anisotropy method for detection of lead (II) ion by a G-quadruplex-inducible DNA aptamer

Zhang

Meng

et al. 2014

Analytica Chimica Acta

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h i g h l i g h t s• A fluorescence anisotropy approach for detection of Pb 2+ was developed.• The strategy was based on bindinginduced allosteric conformational change of aptamer probe.• The sensing mechanism was established by testing the photoinduced electron transfer interaction. in homogeneous solution by a G-rich thrombin binding aptamer (TBA). The TBA labeled with 6-carboxytetramethylrhodamine (TMR) at the seventh thymine nucleotide was used as a fluorescent probe for signaling Pb 2+ . It was found that the aptamer probe had a high FA in the absence of Pb 2+ . This is because the rotation of TMR is restricted by intramolecular interaction with the adjacent guanine bases, which results in photoinduced electron transfer (PET). When the aptamer probe binds to Pb 2+ to form G-quadruplex, the intramolecular interaction should be eliminated, resulting in faster rotation of the fluorophore TMR in solution. Therefore, FA of aptamer probe is expected to decrease significantly upon binding to Pb 2+ . Indeed, we observed a decrease in FA of aptamer probe upon Pb 2+ binding. Circular dichroism, fluorescence spectra, and fluorescence lifetime measurement were used to verify the reliability and reasonability of the sensing mechanism. By monitoring the FA change of the aptamer probe, we were able to real-time detect binding between the TBA probe and Pb 2+ . Moreover, the aptamer probe was exploited as a recognition element for quantification of Pb 2+ in homogeneous solution. The change in FA showed a linear response to Pb 2+ from 10 nM to 2.0 M, with 1.0 nM limit of detection. In addition, this sensing system exhibited good selectivity for Pb 2+ over other metal ions. The method is simple, quick and inherits the advantages of aptamer and FA.

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Mutation Detection in DNA Oligonucleotides Based on a Guanine Quenching Method Coupled with Enzymatic Digestion of Single-Stranded DNA

Cited by 22 publications

References 18 publications

Selection of Peptides Targeting Helix 31 of Bacterial 16S Ribosomal RNA by Screening M13 Phage-Display Libraries

Selection of Peptides Targeting Helix 31 of Bacterial 16S Ribosomal RNA by Screening M13 Phage-Display Libraries

DNA Hybridization in Thermoresponsive Polymer Nanoparticles

A sensitive fluorescence anisotropy method for detection of lead (II) ion by a G-quadruplex-inducible DNA aptamer

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