Detection of a Specific DNA Fragment by Free-Solution Capillary Electrophoresis Using a Fluorescent DNA Probe Measured by Semiconductor Laser-Induced Fluorometry

Takashi Kaneta, Tadashi Okamoto, Totaro Imasaka

Research output: Contribution to journalArticlepeer-review

10 Citations (Scopus)

Abstract

Semiconductor laser-induced fluorescence detection of native (target) DNA has been accomplished by free-solution capillary electrophoresis (CE) using an oligonucleotide labeled with a cyanine dye as a DNA probe. A mixture of probe DNA and target DNA, which is complementary to probe DNA, is incubated to form hybrid DNA; the mixture is then injected into a capillary. Single-stranded probe DNA and double-stranded hybrid DNA were separated by free-solution CE. A 19mer oligonucleotide labeled with fluorescein isothiocyanate (FITC) was used as probe DNA in a preliminary study. The stability of double-stranded DNA during migration was evaluated by changing the temperature of the solution in the capillary, i.e. the applied voltage in CE. The dissociation of double-stranded DNA was appreciable at high applied voltages. Thus, suppressing the temperature in the capillary, i.e. optimization of the voltage, was required in order to prevent the dissociation of double-stranded DNA. Furthermore, a new labeling reagent, a cyanine derivative, was synthesized in order to be applied to semiconductor laser-induced fluorometry. The detection limit was 8×10 -9 M for probe DNA.

Original languageEnglish
Pages (from-to)875-880
Number of pages6
JournalAnalytical Sciences
Volume12
Issue number6
DOIs
Publication statusPublished - 1996

Keywords

  • Capillary electrophoresis
  • Cyanine dye
  • DNA
  • Hybridization
  • Semiconductor laser-induced fluorometry

ASJC Scopus subject areas

  • Analytical Chemistry

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