Method for label-free detection of hybridized DNA targets
AssigneeCorning Incorporated
InventorsAnthony Glenn Frutos, Joydeep Lahiri, Santona Pal, Mark A. Quesada
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Abstract
A method for the detection and analysis of genetic polymorphisms using arrays that do not require labeling of a target nucleic acid sequence. Hybridization of a perfectly complementary nucleic acid target sequence to an oligonucleotide probe sequence results in a displacement and complete removal of a hybridized probe sequence from the same oligonucleotide probe sequence by means of a thermo-kinetic reaction. The removal of the hybridized probe sequence, having a quencher element, increases the intensity of emission by an emitter element on the oligonucleotide probe sequence.
Claims34
A method for detecting or identifying genetic polymorphisms, said method comprising:
- providing (1) a set of oligonucleotide probes, each of said oligonucleotide probes (i) having a nucleic acid sequence that is attached to a solid support, and (ii) differing from each other in at least one base location, and (2) a set of hybridized probes, each of said hybridized probes having a complementary nucleic acid sequence to an oligonucleotide probe, and each of said nucleic acid sequences of said hybridized probes incorporating at least one artificial mismatch site;
- labeling each oligonucleotide probe with an emitter element, and each hybridized probe with a quencher element;
- hybridizing an unlabeled sample nucleic acid target to said oligionucleotide probe, wherein said sample nucleic acid target displaces said hybridized probe from said oligonucleotide probe, wherein said emitter element increases in emission intensity with such displacement.
A method for detecting genetic polymorphisms, said method comprising:
- providing a plurality of DNA duplexes, each having a first nucleic acid sequence that is immobilized to a solid support and labeled with a fluorophore, and a complementary, second nucleic acid sequence that incorporates at least one mismatch site and labeled with a fluorescence quencher;
- exposing said solid support, having said first nucleic acid sequence immobilized thereon, to an unlabeled sample DNA sequence, wherein said sample DNA sequence preferentially dislodges from said first nucleic acid sequence, to which said sample DNA sequence is perfectly complementary, a corresponding second nucleic acid sequence in said DNA duplex, removing said quencher.
A method for detecting genetic polymorphisms, said method comprising:
- immobilizing on a solid support a plurality of first oligonucleotide probes, each of said oligonucleotide probes having a first nucleic acid sequence to which an emitting element is attached;
- providing (1) an unlabeled sample nucleic acid target, and (2) a second probe having a second nucleic acid sequence, said second nucleic acid sequence being a hybridizable, complementary sequence to said first nucleic acid sequence, said second nucleic acid sequence having at least one artificial mismatch site and a quenching element attached to said second nucleic acid sequence;
- hybridzing said second probe and said sample nucleic acid target to said first oligonucleotide, wherein said second probe binds to said first oligonucleotide probe, when said sample nucleic acid target is not complementary, and said quenching element reduces an emission from said emitting element.
An article for detecting or identifying genetic polymorphisms, said article comprises:
- (1) a set of oligonucleotide probes, each of said oligonucleotide probes (i) having a nucleic acid sequence that is immobilized to a solid support and (ii) differing from each other in at least one base location and (2) a set of hybridized probes, each of said hybridized probes having a complementary nucleic acid sequence to an oligonucleotide probe, and each of said nucleic acid sequences of said hybridized probes incorporating at least one artificial mismatch site, each oligonucleotide probe is labeled with an emitter element, and each hybridized probe is labeled with a quencher element, said hybridized probe with said quencher element being displaceable by an unlabeled sample nucleic acid target having a sequence that is perfectly complementary to said oligionucleotide probe, and that said emitter element increases emissions with such displacement.
A kit suitable for performing an assay that detects the presence, absence, or amount of a sample target nucleic acid sequence, the kit comprises:
- an article, said article having a solid support upon which a plurality of first oligonucleotide probes is immobilized, each of said oligonucleotide probes having a first nucleic acid sequence to which an emitter element is attached.
Dependencies are inferred from the claim text.