Patent ID: 7700324
Filing Date: 2010-04-20
Classification: C12Q

Abstract:
1. A method for detecting the presence of a methylated CpG sequence of a specific gene in a sample containing nucleic acids, comprising: a) digesting the sample with a methylation sensitive restriction endonuclease that cleaves only unmethylated CpG sites and generates nucleic acid fragments with blunt ends, and forming cleaved nucleic acids; b) thereafter digesting the cleaved nucleic acids with an isoschizomer of the methylation sensitive restriction endonuclease that cleaves both methylated and unmethylated CpG sites and generates nucleic acid fragments having ends with an overhang, and forming isoschizomer cleaved nucleic acids; c) ligating adaptors to the ends with an overhang of the isoschizomer cleaved nucleic acids and forming ligated nucleic acids, wherein the adaptors have a single stranded overhang complementary to the overhang of each end of the isoschizomer cleaved nucleic acids; and d) amplifying the ligated nucleic acids of step c) with primers that are complementary to the sequences of the adaptors and forming amplified nucleic acids; e) contacting the amplified nucleic acids of step d) with a nucleic acid probe that selectively hybridizes to the gene, wherein the presence of a hybridization of the probe to the amplified nucleic acids indicates the presence of the methylated CpG sequence of the gene in the sample.