Development of the Handy Bio-Strand and its application to genotyping of OPRM1 (A118G).
We previously developed a three-dimensional microarray system, the Bio-Strand, which exhibits advantages in automated DNA analysis in combination with our Magtration Technology. In the current study, we have developed a compact system for the Bio-Strand, the Handy Bio-Strand, which consists of several tools for the preparation of Bio-Strand Tip, hybridization, and detection. Using the Handy Bio-Strand, we performed single nucleotide polymorphism (SNP) genotyping of OPRM1 (A118G) by allele-specific oligonucleotide competitive hybridization (ASOCH). DNA fragments containing SNP sites were amplified from genomic DNA by PCR and then were fixed on a microporous nylon thread. Thus, prepared Bio-Strand Tip was hybridized with allele-specific Cy5 probes (<15mer), on which the SNP site was designed to be located in the center. By optimizing the amount of competitors, the selectivity of Cy5 probes increased without a drastic signal decrease. OPRM1 (A118G) genotypes of 23 human genomes prepared from whole blood samples were determined by ASOCH using the Handy Bio-Strand. The results were perfectly consistent with those determined by PCR direct sequencing. ASOCH using the Handy Bio-Strand would be a very simple and reliable method for SNP genotyping for small laboratories and hospitals.
Ginya
Harumi
H
Precision System Science, Kamihongou, Matsudo, Chiba 271-0064, Japan. harumi.ginya@pss.co.jp
Asahina
Junko
J
Yoshida
Mamiko
M
Segawa
Osamu
O
Asano
Tsutomu
T
Ikeda
Hideo
H
Hatano
Yukiko Miyashita
YM
Shishido
Madoka
M
Johansson
Britt-Marie
BM
Zhou
Qin
Q
Hallberg
Mathias
M
Takahashi
Masaaki
M
Nyberg
Fred
F
Tajima
Hideji
H
Yohda
Masafumi
M
D000483:Alleles; D001483:Base Sequence; D004247:DNA / Q000235:genetics / Q000302:isolation & purification; D004867:Equipment Design; D005838:Genotype; D006801:Humans; D020411:Oligonucleotide Array Sequence Analysis / Q000295:instrumentation* / Q000379:methods; D015345:Oligonucleotide Probes / Q000235:genetics; D020641:Polymorphism, Single Nucleotide; D017450:Receptors, Opioid, mu / Q000235:genetics*
C485764:OPRM1 protein, human; D015345:Oligonucleotide Probes; D017450:Receptors, Opioid, mu; D004247:DNA