Denaturing gradient-based two-dimensional gene mutation scanning in a polymer microfluidic network

Jesse S. Buch, Frederick Rosenberger, W Edward Jr. Highsmith, Christopher Kimball, Don L. DeVoe, Cheng S. Lee

Research output: Contribution to journalArticle

21 Citations (Scopus)

Abstract

An integrated two-dimensional (2-D) DNA separation platform, combining standard gel electrophoresis with temperature gradient gel electrophoresis (TGGE) on a polymer microfluidic chip, is reported. Rather than sequentially sampling DNA fragments eluted from standard gel electrophoresis, size-resolved fragments are simultaneously electrokinetically transferred into an array of orthogonal microchannels and screened for the presence of sequence heterogeneity by TGGE in a parallel and high throughput format. A bulk heater assembly is designed and employed to externally generate a temporal temperature gradient along an array of TGGE channels. Extensive finite element modeling is performed to determine the optimal geometries of the microfluidic network for minimizing analyte band dispersion caused by interconnected channels in the network. A pH-mediated on-chip analyte stacking strategy is employed prior to the parallel TGGE separations to further reduce additional band broadening acquired during the electrokinetic transfer of DNA fragments between the first and second separation dimensions. A comprehensive 2-D DNA separation is completed in less than 5 min for positive detection of single-nucleotide polymorphisms in multiplex PCR products that vary in size and sequence.

Original languageEnglish (US)
Pages (from-to)392-400
Number of pages9
JournalLab on a Chip - Miniaturisation for Chemistry and Biology
Volume5
Issue number4
DOIs
StatePublished - Apr 2005

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Denaturing Gradient Gel Electrophoresis
Microfluidics
Electrophoresis
Polymers
Gels
Thermal gradients
Genes
Scanning
Mutation
DNA
Multiplex Polymerase Chain Reaction
Single Nucleotide Polymorphism
Nucleotides
Microchannels
Polymorphism
Temperature
Throughput
Sampling
Geometry

ASJC Scopus subject areas

  • Clinical Biochemistry

Cite this

Denaturing gradient-based two-dimensional gene mutation scanning in a polymer microfluidic network. / Buch, Jesse S.; Rosenberger, Frederick; Highsmith, W Edward Jr.; Kimball, Christopher; DeVoe, Don L.; Lee, Cheng S.

In: Lab on a Chip - Miniaturisation for Chemistry and Biology, Vol. 5, No. 4, 04.2005, p. 392-400.

Research output: Contribution to journalArticle

Buch, Jesse S. ; Rosenberger, Frederick ; Highsmith, W Edward Jr. ; Kimball, Christopher ; DeVoe, Don L. ; Lee, Cheng S. / Denaturing gradient-based two-dimensional gene mutation scanning in a polymer microfluidic network. In: Lab on a Chip - Miniaturisation for Chemistry and Biology. 2005 ; Vol. 5, No. 4. pp. 392-400.
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