The effects of mismatches on hybridization in DNA microarrays: determination of nearest neighbor parameters

Hooyberghs, Jef; Hummelen, Paul Van; Carlon, Enrico

doi:10.1093/nar/gkp109

Cited by 58 publications

(85 citation statements)

References 20 publications

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“…the subtraction from the hybridization free energy of the perfect match probe, which cannot be determined from the fitting procedure. 20 The similarities of the plots in both columns of Fig. 3 show a correlation between ∆G sol and ∆G µarray .…”

Section: Custom Agilent Arraysmentioning

confidence: 72%

“…3 show a correlation between ∆G sol and ∆G µarray . 20 However, when plotted as a function of ∆∆G µarray the data "collapse" better into single master curves, compared to the plots as a function of the nearest-neighbor model parameters in solution which are more spread. Figures 3(a,b) show the measured intensities for a target strand of length L = 30 hybridizing for 17h, which is the typical hybridization time of the standard protocol used in biological experiments.…”

Section: Custom Agilent Arraysmentioning

confidence: 99%

“…We review some experimental results recently obtained by two of us 16,20 Here, ∆G sol is calculated based on nearest neighbor parameters by SantaLucia 19 at 1M [Na+] concentration. The hybridization buffer in the experiment is the standard Agilent buffer which contains a buffering agent, monovalent cation and a ionic surfactant.…”

Section: Custom Agilent Arraysmentioning

confidence: 99%

“…The ∆G µarrays are obtained from a linear least-square fit of the data, as explained in Ref. 20 The left column of Fig. 3 plots the data as function of ∆∆G µarray = ∆G µarray − ∆G PMµarray , i.e.…”

Section: Custom Agilent Arraysmentioning

confidence: 99%

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Nonequilibrium Effects in DNA Microarrays: A Multiplatform Study

et al. 2011

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It has recently been shown that in some DNA microarrays the time needed to reach thermal equilibrium may largely exceed the typical experimental time, which is about 15h in standard protocols (Hooyberghs et al. Phys. Rev. E 81, 012901 (2010)). In this paper we discuss how this breakdown of thermodynamic equilibrium could be detected in microarray experiments without resorting to real time hybridization data, which are difficult to implement in standard experimental conditions. The method is based on the analysis of the distribution of fluorescence intensities I from different spots for probes carrying base mismatches. In thermal equilibrium and at sufficiently low concentrations, log I is expected to be linearly related to the hybridization free energy ∆G with a slope equal to 1/RT exp , where T exp is the experimental temperature and R is the gas constant. The breakdown of equilibrium results in the deviation from this law. A model for hybridization kinetics explaining the observed experimental behavior is discussed, the so-called 3-state model. It predicts that deviations from equilibrium yield a proportionality of log I to ∆G/RT eff . Here, T eff is an "effective" temperature, higher than the experimental one. This behavior is indeed observed in some experiments on Agilent arrays. 16,20 We analyze experimental data from two other microarray platforms and discuss, on the basis of the results, the attainment of equilibrium in these cases. Interestingly, the same 3-state model predicts a (dynamical) saturation of the signal at values below the expected one at equilibrium.

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Section: Custom Agilent Arraysmentioning

confidence: 72%

Section: Custom Agilent Arraysmentioning

confidence: 99%

Section: Custom Agilent Arraysmentioning

confidence: 99%

Section: Custom Agilent Arraysmentioning

confidence: 99%

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Nonequilibrium Effects in DNA Microarrays: A Multiplatform Study

et al. 2011

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“…When nucleic acid targets hybridize on immobilized surface probes its equilibrium state can be described by the Langmuir isotherm [20,21,22]. Assuming that the fraction of hybridized probes in a microarray spot is proportional to the measured signal intensity, for a DNA double helix in thermodynamic equilibrium, the isotherm can be written as…”

Section: Langmuir Isotherm and Dynamic Rangementioning

confidence: 99%

Dynamic range extension of hybridization sensors

Hadiwikarta

Carlon

Hooyberghs

2015

Biosensors and Bioelectronics

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In hybridization based nucleic acid sensors the stringency of hybridization poses a challenge to design and experiment. For a given set of experimental parameters the affinity window of probe-target interaction is always limited and vice versa for a given probe set design, changes in experimental conditions can easily bring some measurements out of detection range. In this paper we introduce and apply a strategy to extend this dynamic range for affinity sensors, sensors which measure the amount of hybridized molecules after equilibrium is reached. The method relies on concepts of additivity of nucleic acids hybridization free energies and on equilibrium isotherms. It consists in combining the measurements from probes with different length, by appropriately rescaling the measured signals. We test the validity of the approach on experiments and show that by combining probes with hybridizing regions of length 21, 23 and 25 nucleotides we manage to extend the dynamic range of the intensity signals by a factor 25. The presented concept is easy to extend, platform free and applies to any hybridization based affinity sensor.

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Target concentration dependence of DNA melting temperature on oligonucleotide microarrays

Ozel

Srivannavit

Rouillard

et al. 2012

Biotechnology Progress

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The design of microarrays is currently based on studies focusing on DNA hybridization reaction in bulk solution. However, the presence of a surface to which the probe strand is attached can make the solution-based approximations invalid, resulting in sub-optimum hybridization conditions. To determine the effect of surfaces on DNA duplex formation, the authors studied the dependence of DNA melting temperature (T(m)) on target concentration. An automated system was developed to capture the melting profiles of a 25-mer perfect-match probe-target pair initially hybridized at 23°C. Target concentrations ranged from 0.0165 to 15 nM with different probe amounts (0.03-0.82 pmol on a surface area of 10(18) Å(2)), a constant probe density (5 × 10(12) molecules/cm(2)) and spacer length (15 dT). The authors found that T(m) for duplexes anchored to a surface is lower than in-solution, and this difference increases with increasing target concentration. In a representative set, a target concentration increase from 0.5 to 15 nM with 0.82 pmol of probe on the surface resulted in a T(m) decrease of 6°C when compared with a 4°C increase in solution. At very low target concentrations, a multi-melting process was observed in low temperature domains of the curves. This was attributed to the presence of truncated or mismatch probes.

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The effects of mismatches on hybridization in DNA microarrays: determination of nearest neighbor parameters

Cited by 58 publications

References 20 publications

Nonequilibrium Effects in DNA Microarrays: A Multiplatform Study

Nonequilibrium Effects in DNA Microarrays: A Multiplatform Study

Dynamic range extension of hybridization sensors

Target concentration dependence of DNA melting temperature on oligonucleotide microarrays

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