Isolation of CpG islands by thermal segregation and enzymatic selection-amplification method

A method of amplifying a plurality of GC-rich DNA molecules without cloning, comprising:

• providing a plurality of DNA molecules, said plurality having molecules comprising one or more regions that are GC-poor and having molecules comprising one or more regions that are GC-rich;
• subjecting the plurality of DNA molecules to denaturation and renaturation conditions to substantially denature GC-poor regions but not to denature GC-rich regions, wherein the GC-rich regions are referred to as thermo-resistant DNA molecules;
• attaching an adaptor by ligation to ends of the thermo-resistant DNA molecules to produce adaptor-linked thermo-resistant DNA molecules;
• and
• performing one of the following without cloning:
• (a) subjecting the adaptor-linked thermo-resistant DNA molecules to one or more methylation-sensitive restriction enzymes, thereby cleaving non-methylated DNA and retaining methylated DNA fragments to produce methylated GC-rich DNA fragments;
• and
• PCR amplifying the methylated GC-rich DNA fragments;
• or
• (b) subjecting the adaptor-linked thermo-resistant DNA molecules to one or more methylation-specific restriction enzymes, thereby cleaving methylated DNA and retaining non-methylated DNA fragments to produce non-methylated GC-rich DNA fragments;
• and
• PCR amplifying the non-methylated GC-rich DNA fragments.