Nucleic acid detection assay

A method for determining the methylation status of a potential methylation site on genomic nucleic acid comprising:

• treating genomic nucleic acid with an agent which modifies cytosine bases but does not modify 5-methyl-cytosine bases under conditions to form a modified nucleic acid template from two complementary strands of genomic nucleic acid containing a potential methylation site;
• providing a first clamp containing a first capture sequence and a second capture sequence, first capture sequence being complementary to a region flanking one side of the potential methylation site in the first strand of the modified nucleic acid template, and the second capture sequence being complementary to a region flanking one side of the potential methylation site in the second strand of the modified nucleic acid template;
• providing a second clamp containing a third capture sequence and a fourth capture sequence, the third capture sequence being complementary to a region flanking the other side the potential methylation site in the first strand of the modified nucleic acid template, and the fourth capture sequence being complementary to a region flanking the other side of the potential methylation site in the second strand of the modified nucleic acid template;
• allowing the first clamp and the second clamp to hybridise to the two complementary strands of the modified nucleic acid template;
• ligating the hybridised clamps to form a circular probe spanning the potential methylation site in the complementary strands of the modified nucleic acid template;
• digesting the modified nucleic acid template to obtain the circular probe;
• and
• detecting the circular probe and determining the methylation status of the potential methylation site in the modified genomic nucleic acid.