Patent ID: 8728733
Filing Date: 2014-05-20
Classification: C12Q

Abstract:
1. A method for identifying methyl CpG island-associated genome signature tags comprising: a) providing a sample containing genomic DNA; b) isolating the genomic DNA from the sample; c) contacting the isolated DNA from step b) with a fragmenting type II restriction enzyme, said fragmenting restriction enzyme having a low probability of cleaving G-C rich DNA sequences, under conditions appropriate for substantially complete digestion of the DNA, thereby generating a plurality of DNA fragment species, each having complementary cohesive termini; d) separating methylated CpG island (Me-CpGI) fragments from fragments in which CpG sequences are not methylated by contacting the DNA fragment species of step c) with an affinity resin that specifically recognizes and binds Me-CpG sequences thereby capturing the Me-CpGI fragments on the resin; e) incubating the captured Me-CpGI fragments with a molar excess of duplex linker having a type IIS restriction enzyme recognition sequence and one cohesive terminus compatible with termini generated by the fragmenting restriction enzyme, under conditions appropriate for ligating one duplex linker to each terminus of the captured fragments of step d); f) contacting the ligation products of step e) with the type IIS restriction enzyme, under conditions appropriate for complete digestion of the captured Me-CpGI fragments, thereby releasing the duplex linkers and appended Me-CpGI-associated signature tags; g) recovering the released linkers and appended tags; h) incubating the recovered linkers and tags of step i) recovering the ligation products of step h); j) determining the nucleotide sequence of a statistically significant number of appended signature tags to generate a listing of methyl CpG Island-associated signature tags, thereby identifying methyl CpG Island-associated genome signature tags.