title: Quantitative analysis of aberrant p16 methylation using real-time quantitative methylation-specific polymerase chain reaction.
abstract: We have developed a quantitative method for methylation analysis of the p16 gene based on real-time methylation-specific PCR (MSP). Real-time MSP is sensitive enough to detect down to 10 genome equivalents of the methylated p16 sequence. Application of real-time MSP to DNA from tumor-derived cell lines revealed complete concordance with conventional MSP analysis. Quantitative data generated by real-time MSP were expressed as the methylation index, which was defined as the percentage of bisulfite-converted DNA that consisted of methylated target sequences. The methylation index was shown to be inversely correlated with p16 gene transcription during demethylation treatment of cell lines with 5-aza-2'-deoxycytidine. The application of real-time MSP to bone marrow aspirates from patients with multiple myeloma revealed complete concordance with conventional MSP analysis. Real-time quantitative MSP may have applications in elucidating diverse biological processes involving DNA methylation and may become a valuable diagnostic tool for detecting tumor-associated epigenetic changes in cancer patients.
authors.affiliation: Department of Chemical Pathology, The Chinese University of Hong Kong, Prince of Wales Hospital, Shatin, New Territories. loym@cuhk.edu.hk
authors.forename: Y M
authors.initials: YM
authors.lastname: Lo
authors.forename: I H
authors.initials: IH
authors.lastname: Wong
authors.forename: J
authors.initials: J
authors.lastname: Zhang
authors.forename: M S
authors.initials: MS
authors.lastname: Tein
authors.forename: M H
authors.initials: MH
authors.lastname: Ng
authors.forename: N M
authors.initials: NM
authors.lastname: Hjelm
mesh_terms: D019175:DNA Methylation; D019942:Genes, p16; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D013997:Time Factors