The Effect of Temperature Oscillations on DNA Sequencing by Capillary Electrophoresis

Voss, Karl O.; Roos, H. Pieter; Dovic̀hi, Norman J.

doi:10.1021/ac000977m

Cited by 16 publications

(12 citation statements)

References 16 publications

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“…We do not have a convincing hypothesis to explain these variations at the present time, however one potential cause is the temperature variations in the nonthermostated capillary. Small local changes in temperature may result in buffer viscosity changes, which previously have been shown to affect separations [21].…”

Section: Electropherograms Of Individually Detected Microspheresmentioning

confidence: 98%

Determination of individual microsphere properties by capillary electrophoresis with laser-induced fluorescence detection

2002

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Capillary electrophoresis with postcolumn laser-induced fluorescence detection was used to individually detect 6.0, 1.0, 0.5, and 0.2 num diameter polystyrene microspheres and individually measure their electrophoretic mobility. The analysis of a nanoliter-size volume from a microsphere suspension results in an electropherogram characterized by several narrow spikes in a well-defined migration time window. Each spike is associated with one microsphere because, when one single microsphere is introduced into the capillary by micromanipulation, the electropherogram has only one spike in the same migration time window. The distributions of individual measurements resulting from an electropherogram were used to evaluate the reproducibility from run to run, observe the effect of sodium dodecyl sulfate (SDS) added to the running buffer, and to investigate the origin of electrophoretic dispersion. As expected from the interactions between microspheres and SDS, the addition of this surfactant to the running buffer narrowed the range and shifted the average electrophoretic mobility to more negative values. After evaluating common sources of broadening in capillary electrophoresis, electrophoretic dispersion was attributed to microsphere heterogeneity. Unlike electropherograms displaying Gaussian-like profiles, the two-dimensional representations of the individual measurements provide a new alternative to evaluate and study electrophoretic-related properties of microspheres.

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Section: Electropherograms Of Individually Detected Microspheresmentioning

confidence: 98%

Determination of individual microsphere properties by capillary electrophoresis with laser-induced fluorescence detection

2002

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“…The Monte Carlo method can be used for modeling the diffusion processes without "explicitly" solving System 5 [22]. This method, however, requires longer calculation times to achieve the same accuracy as the explicit numerical solution of System 5.…”

Section: Numerical Solutionmentioning

confidence: 99%

Transverse diffusion of laminar flow profiles – a generic method for mixing reactants in capillary microreactor

Krylova

Okhonin

Krylov³

2009

J of Separation Science

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The capillary is an attractive format for integrated microanalyses, which start with the injection of separate reactants into the capillary and their mixing inside the capillary. Due to the nonturbulent nature of flow inside the capillary, mixing reactants in a generic way is a challenging task. Three approaches have been suggested as a solution: mixing by electrophoresis, mixing by longitudinal diffusion, and, most recently, mixing by transverse diffusion of laminar flow profiles (TDLFP). This is the first review on TDLFP, describing: (i) the physical basis of the method, (ii) its theory, (iii) analytical and numerical solutions for the calculation of concentration profiles of mixed reactants, (iv) up-to-date applications, and (v) problems to be solved and future directions.

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“…It was observed that a temperature change as small as 0.058C was sufficient to induce an observable mobility shift, due to the change of equilibrium between partially melted and double stranded DNA molecules (unpublished data). In addition, it was observed that minute oscillations as small as l0.18C in a capillary's temperature resulted in significant degradation in the number of theoretical plates due to the laminar flow induced by the periodic thermal expansion and contraction of the separation medium [41]. Therefore, it is crucial that accuracy and precision better than l0.058C be maintained to ensure reproducibility in the CDCE process, if isothermal temperature control is employed.…”

Section: Resultsmentioning

confidence: 99%

Design of an automated multicapillary instrument with fraction collection for DNA mutation discovery by constant denaturant capillary electrophoresis (CDCE)

Deka²,

Glassner³

et al. 2005

J of Separation Science

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A fundamental goal ingenomics is the discovery of genetic variation that contributes to disease states or to differential drug responses. Single nucleotide polymorphism (SNP) detection has been the focus of much attention in the study of genetic variation over the last decade. These SNPs typically occur at a frequency greater than 1% in the human genome. Recently, low-frequency alleles are also being increasingly recognized as critical to obtain an improved understanding of the correlation between genetic variation and disease. Although many methods have been reported for the discovery and scoringof SNPs, sensitive, automated, and cost-effective methods and platforms for the discovery of low-frequency alleles are not yet readily available. We describe here an automated multicapillary instrument for high-throughput detection of low-frequency alleles from pooled samples using constant denaturant capillary electrophoresis. The instrument features high optical sensitivity (1 x 10(-12) M fluorescein detection limit), precise and stable temperature control (+/- 0.01degrees C), and automation for sample delivery, injection, matrix replacement, and fraction collection. The capillary array is divided into six groups of four capillaries, each of which can be independently set at any temperature ranging from room temperature to 90 degrees C. The key performance characteristics of the instrument are reported.

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The Effect of Temperature Oscillations on DNA Sequencing by Capillary Electrophoresis

Cited by 16 publications

References 16 publications

Determination of individual microsphere properties by capillary electrophoresis with laser-induced fluorescence detection

Determination of individual microsphere properties by capillary electrophoresis with laser-induced fluorescence detection

Transverse diffusion of laminar flow profiles – a generic method for mixing reactants in capillary microreactor

Design of an automated multicapillary instrument with fraction collection for DNA mutation discovery by constant denaturant capillary electrophoresis (CDCE)

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