MagSNiPer: a new single nucleotide polymorphism typing method based on single base extension, magnetic separation, and chemiluminescence.
We have developed a new method for typing single nucleotide polymorphisms (SNPs), MagSNiPer, based on single base extension, magnetic separation, and chemiluminescence. Single base nucleotide extension reaction is performed with a biotinylated primer whose 3' terminus is contiguous to the SNP site with a tag-labeled ddNTP. Then the primers are captured by magnetic-coated beads with streptavidin, and unincorporated labeled ddNTP is removed by magnetic separation. The magnetic beads are incubated with anti-tag antibody conjugated with alkaline phosphatase. After the removal of excess conjugates by magnetic separation, SNP typing is performed by measuring chemiluminescence. The incorporation of labeled ddNTP is monitored by chemiluminescence induced by alkaline phosphatase. MagSNiPer is a simple and robust SNP typing method with a wide dynamic range and high sensitivity. Using MagSNiPer, we could perform SNP typing with as little as 10(-17) mol of template DNA.
Kakihara
Fumiko
F
Department of Biotechnology and Life Science, Tokyo University of Agriculture and Technology, 2-24-16, Naka-cho, Koganei, Tokyo 184-8588, Japan.
Kurebayashi
Yukari
Y
Tojo
Yuriko
Y
Tajima
Hideji
H
Hasegawa
Setsuo
S
Yohda
Masafumi
M
D000328:Adult; D019389:Cytochrome P-450 CYP2D6 / Q000032:analysis / Q000235:genetics; D005819:Genetic Markers; D006801:Humans; D008163:Luminescent Measurements; D020641:Polymorphism, Single Nucleotide; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA / Q000379:methods
D005819:Genetic Markers; D019389:Cytochrome P-450 CYP2D6