Single-nucleotide polymorphism analysis using fluorescence resonance energy transfer between DNA-labeling fluorophore, fluorescein isothiocyanate, and DNA intercalator, POPO-3, on bacterial magnetic particles.
To develop an analytical system for single-nucleotide polymorphisms (SNPs), the fluorescence resonance energy transfer (FRET) technique was employed on a bacterial magnetic particle (BMP) surface. A combination of fluorescein isothiocyanate (FITC; excitation 490 nm/emission 520 nm) labeled at the 5' end of DNA and an intercalating compound (POPO-3, excitation 534 nm/emission 570 nm) was used to avoid the interference from light scattering caused by nanoparticles. After hybridization between target DNA immobilized onto BMPs and FITC-labeled probes, fluorescence from POPO-3, which was excited by the energy from the FITC, was detected. The major homozygous (ALDH2*1), heterozygous (ALDH2*1/*2), and minor homozygous (ALDH2*2) genotypes in the blood samples were discriminated by this method. The assay described herein allows for a simple and rapid SNP analysis using a fully automated system.
Nakayama
Hideki
H
Department of Biotechnology, Tokyo University of Agriculture and Technology, 2-24-16, Koganei, Tokyo 184-8588, Japan.
Arakaki
Atsushi
A
Maruyama
Kohei
K
Takeyama
Haruko
H
Matsunaga
Tadashi
T
D001426:Bacterial Proteins; D001583:Benzoxazoles; D004247:DNA / Q000032:analysis* / Q000235:genetics*; D016650:Fluorescein-5-isothiocyanate; D031541:Fluorescence Resonance Energy Transfer / Q000379:methods*; D005456:Fluorescent Dyes; D020869:Gene Expression Profiling / Q000379:methods*; D007364:Intercalating Agents; D008565:Membrane Proteins; D036103:Nanotechnology / Q000379:methods; D043942:Nanotubes; D020641:Polymorphism, Single Nucleotide / Q000235:genetics*; D013194:Staining and Labeling / Q000379:methods
D001426:Bacterial Proteins; D001583:Benzoxazoles; D005456:Fluorescent Dyes; D007364:Intercalating Agents; D008565:Membrane Proteins; C405425:MpsA protein, Magnetospirillum sp.; C120457:POPO 3; D004247:DNA; D016650:Fluorescein-5-isothiocyanate