Methods for Detecting and Measuring Specific Nucleic Acid Sequences

A method for detecting the presence or absence of a target sequence, comprising the steps of:

• hybridizing an oligonucleotide to a nucleic acid sample and a fluorophore-labeled reporter sequence, wherein the oligonucleotide comprises (1) a hairpin-forming sequence capable of forming a stem-loop and (2) a sequence capable of hybridizing under nucleic acid array hybridization conditions to the fluorophore-labeled reporter sequence, wherein the oligonucleotide is capable of hybridization under nucleic acid array hybridization conditions to the target sequence, and hybridization of the oligonucleotide to the target sequence prevents formation of the stem-loop in the oligonucleotide, and wherein formation of the stem-loop quenches fluorescence signals of the fluorophore-labeled reporter sequence when the reporter sequence is hybridized to the oligonucleotide;
• and
• detecting the fluorescent signals of the reporter sequence,
• wherein an increase in fluorescence signals of the fluorophore-labeled reporter sequence in the presence of the nucleic acid sample compared to that in the absence of the nucleic acid sample is suggestive of the presence of the target sequence in the sample, and no significant change in fluorescence signals of the fluorophore-labeled reporter sequence in the presence of the nucleic acid sample compared to that in the absence of the nucleic acid sample is suggestive of the absence of the target sequence in the sample.

A method for detecting a sequence difference between a target sequence and a sequence of interest, comprising the steps of:

• hybridizing an oligonucleotide to the sequence of interest and a fluorophore-labeled reporter sequence, wherein the oligonucleotide comprises (1) a hairpin-forming sequence capable of forming a stem-loop and (2) a sequence capable of hybridizing under nucleic acid array hybridization conditions to the fluorophore-labeled reporter sequence, wherein the oligonucleotide comprises a sequence that is complementary to the target sequence, and hybridization of the target sequence to the oligonucleotide prevents formation of the stem-loop in the oligonucleotide, and wherein formation of the stem-loop quenches fluorescence signals of the fluorophore-labeled reporter sequence when the reporter sequence is hybridized to the oligonucleotide;
• and
• detecting the fluorescent signals of the reporter sequence,
• wherein an decrease in fluorescence signals of the fluorophore-labeled reporter sequence in the presence of the sequence of interest compared to that in the presence of the target sequence, together with an increase in fluorescence signals of the fluorophore-labeled reporter sequence in the presence of the sequence of interest compared to that in the absence of the sequence of interest, is suggestive that the sequence of interest is homologous to but different from the target sequence.