Quantitative DNA methylation analysis by fluorescent polymerase chain reaction single-strand conformation polymorphism using an automated DNA sequencer.
A novel DNA methylation assay technique, termed bisulfite single-strand conformation polymorphism (bisulfite-SSCP), is a combination of sodium-bisulfite modification and fluorescence-based polymerase chain reaction (PCR)-SSCP. After bisulfite treatment followed by PCR amplification, methylated and unmethylated alleles can be simultaneously separated in a nondenaturing gel using an automated DNA sequencer. Using bisulfite-SSCP, methylation of hMLH1 was detected in a quantitative manner. This method is not only simple, quick, accurate, and quantitative, but detailed information about methylation is also available with less work. Methylation analysis of large numbers of samples for multiple loci will be facilitated by bisulfite-SSCP.
Suzuki
H
H
First Department of Internal Medicine, Sapporo Medical University, Japan.
Itoh
F
F
Toyota
M
M
Kikuchi
T
T
Kakiuchi
H
H
Hinoda
Y
Y
Imai
K
K
D048868:Adaptor Proteins, Signal Transducing; D001322:Autoanalysis; D015820:Cadherins / Q000235:genetics; D002352:Carrier Proteins; D002478:Cells, Cultured; D019941:Cyclin-Dependent Kinase Inhibitor p16 / Q000235:genetics; D004247:DNA / Q000032:analysis*; D019175:DNA Methylation; D016147:Genes, Tumor Suppressor; D006801:Humans; D016014:Linear Models; D000070957:MutL Protein Homolog 1; D009363:Neoplasm Proteins / Q000235:genetics; D009687:Nuclear Proteins; D016133:Polymerase Chain Reaction; D018807:Polymorphism, Single-Stranded Conformational; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA; D013447:Sulfites / Q000494:pharmacology; D014407:Tumor Cells, Cultured
D048868:Adaptor Proteins, Signal Transducing; D015820:Cadherins; D002352:Carrier Proteins; D019941:Cyclin-Dependent Kinase Inhibitor p16; C501486:MLH1 protein, human; D009363:Neoplasm Proteins; D009687:Nuclear Proteins; D013447:Sulfites; D004247:DNA; D000070957:MutL Protein Homolog 1; C042345:hydrogen sulfite