Development and evaluation of an automated workstation for single nucleotide polymorphism discrimination using bacterial magnetic particles.
We designed an automated workstation for magnetic particle-based single nucleotide polymorphism (SNP) discrimination of ALDH genotypes. Bacterial magnetic particles (BMPs) extracted from Magnetospirillum magneticum AMB-1 were used as DNA carriers. The principle for SNP discrimination in this study was based on fluorescence resonance energy transfer (FRET) between FITC (donor) and POPO-3 (acceptor) bound to double-stranded DNA. The workstation is equipped with a 96-way automated pipetter which collects and dispenses fluids as it moves in x- and z-directions. The platform contains a disposable tip rack station, a reagent vessel serving as a stock for POPO-3 and FITC-labeled probes and a reaction station for a 96-well microtiter plate. BMPs were collected by attaching a neodymium iron boron sintered (Nd-Fe-B) magnet on the bottom of the microtiter plate. This system permits the simultaneous heating and magnetic separation of 96 samples per assay. The genotypes ALDH2*1 and ALDH2*2 were discriminated by calculating the relative fluorescence intensities on BMPs.
Tanaka
Tsuyoshi
T
Department of Biotechnology, Tokyo University of Agriculture and Technology, 2-24-16 Naka-cho Koganei, Tokyo 184-8588, Japan.
Maruyama
Kohei
K
Yoda
Kiyoushi
K
Nemoto
Etsuo
E
Udagawa
Yuuji
Y
Nakayama
Hideki
H
Takeyama
Haruko
H
Matsunaga
Tadashi
T
D000444:Aldehyde Dehydrogenase / Q000032:analysis / Q000235:genetics*; D000071396:Aldehyde Dehydrogenase, Mitochondrial; D001426:Bacterial Proteins / Q000737:chemistry*; D015342:DNA Probes / Q000737:chemistry*; D004867:Equipment Design; D019544:Equipment Failure Analysis; D031541:Fluorescence Resonance Energy Transfer / Q000295:instrumentation*; D006801:Humans; D008565:Membrane Proteins / Q000737:chemistry*; D008838:Microcomputers; D020411:Oligonucleotide Array Sequence Analysis / Q000295:instrumentation* / Q000379:methods; D020641:Polymorphism, Single Nucleotide / Q000235:genetics*; D015203:Reproducibility of Results; D012371:Robotics / Q000295:instrumentation* / Q000379:methods; D012680:Sensitivity and Specificity; D014584:User-Computer Interface
D001426:Bacterial Proteins; D015342:DNA Probes; D008565:Membrane Proteins; C405425:MpsA protein, Magnetospirillum sp.; C096127:ALDH2 protein, human; D000444:Aldehyde Dehydrogenase; D000071396:Aldehyde Dehydrogenase, Mitochondrial