Patent ID: 8017327

Claim:
A method for detecting a single nucleotide polymorphism in a target nucleic acid comprising: (a) providing (i) a linear or two-dimensional microarray of discrete regions, each having a defined area, formed on an upper surface of a solid support, each region of the microarray comprising a signal probe and an invader probe, wherein the signal probe or the signal and the invader probe are immobilized on the upper surface, and (ii) a plurality of different samples having an amplified target nucleic acid suspected of having a single polynucleotide polymorphism at discrete regions in the microarray, wherein the invader probe comprises sequences selected to anneal to a single-strand of the amplified target nucleic acid 5′ to the single polynucleotide polymorphism and a 3′ most nucleotide that does not anneal to the single polynucleotide polymorphism, wherein the signal probe comprises a fluorescent label linked to sequences selected to not anneal to the single-strand of the amplified target nucleic acid or to the invader probe linked to sequences selected to anneal to the target nucleic acid with a single nucleotide polymorphism, wherein the signal probe further comprises a fluorescence quencher linked to the sequences selected to anneal to the single-strand of the amplified target nucleic acid; (b) annealing at each discrete region in the microarray the invader probe and the signal probe to the single-strand of the amplified target nucleic acid suspected of having a single polynucleotide polymorphism to provide a sample complex; (c) contacting the sample complex with a flap endonuclease to activate a fluorescence response; wherein if the signal probe anneals to the single-strand of the amplified target nucleic acid at a single polymorphic nucleotide, the flap endonuclease cleaves the signal probe 3′ to the single polymorphic nucleotide producing a cleaved 5′ flap sequence that produces the fluorescent response; and (d) detecting the fluorescence response in the discrete regions, wherein fluorescence in a discrete region is indicative of the presence of the single nucleotide polymorphism in the sample.