Nucleic acid detection
AssigneeZoragen, Inc.
InventorNora Szasz
Life of this patent
Estimating the term…
Abstract
The invention relates to methods for the detection of the amount of a nucleic acid in a sample. The described methods exploit the ability to physically pair nucleic acid molecules in a sample that have a reference sequence with nucleic acid molecules in the sample that have a target sequence. The presence of unpaired target or reference sequence following such physical pairing indicates a difference in the amount of the target sequence versus the reference sequence. The methods are broadly applicable for the determination of differences in the amounts of nucleic acids in diagnostic and research applications.
Claims72
A method of determining the amount of a target nucleic acid relative to the amount of a reference nucleic acid in a nucleic acid sample, comprising:
- A) incubating said sample under conditions that permit specific binding of first and second probes to said target and reference nucleic acids, respectively, wherein
- (i) said nucleic acid sample comprises said target and reference nucleic acids;
- (ii) said first probe comprises a target nucleic acid binding sequence and a second probe binding moiety;
- (iii) said second probe comprises a reference nucleic acid binding sequence and a first probe binding moiety;
- B) placing said first and second probes under conditions that permit the pairing of said probes, wherein said pairing comprises binding of said first probe binding moiety to said second probe binding moiety, to form paired probes;
- and
- C) detecting unpaired probe, wherein said detecting is indicative of a difference in the amount of target and reference nucleic acids in said sample.
A method of determining the amount of a target nucleic acid sequence relative to the amount of a reference nucleic acid sequence in a nucleic acid sample, the method comprising:
- (a) providing a sample, said sample comprising said target nucleic acid sequence and said reference nucleic acid sequence;
- (b) contacting said sample, under conditions that permit hybridization, with first and second probes that have the following characteristics:
- (i) said first probe comprises a first binding partner moiety and a sequence that binds specifically to said target nucleic acid sequence, wherein said first binding partner moiety can bind a second binding partner moiety on said second probe when said probes are placed under conditions that permit binding of said binding partner moieties;
- and
- (ii) said second probe comprises a second binding partner moiety and a sequence that binds specifically to said reference nucleic acid sequence;
- (c) placing said first and second probes of step (b) under conditions that permit said first binding partner moiety of said first probe to interact with said second binding partner moiety of said second probe such that said first and second probes become bound to each other to form paired probes;
- and
- (d) detecting a probe that is not bound to another probe, wherein said detecting indicates a difference in the amount of target and reference nucleic acid sequences present in said sample.
A method of determining the amount of a target nucleic acid relative to the amount of a reference nucleic acid in a nucleic acid sample, the method comprising:
- (a) contacting a nucleic acid sample with a probe under conditions that permit hybridization, wherein said probe comprises a sequence that specifically binds to said target nucleic acid sequence and a sequence that specifically binds to said reference nucleic acid sequence;
- (b) detecting unhybridized probe, target or reference nucleic acid, wherein hybridized probe and/or target and/or reference nucleic acid is resistant to detection, and wherein said detecting determines the amount of said target nucleic acid relative to the amount of said reference nucleic acid present in said nucleic acid sample.
A method of detecting a chromosomal abnormality, said method comprising the steps of
- (a) obtaining a nucleic acid sample;
- (b) contacting said sample, under conditions that permit hybridization, with first and second probes that have the following characteristics:
- (i) said first probe comprises a sequence that binds specifically to said target sequence, and a first binding partner moiety, wherein said first binding partner moiety can bind a second binding partner moiety on said second probe, when said probes are placed under conditions that permit binding of said binding partner moieties;
- and
- (ii) said second probe comprises a sequence that binds specifically to said reference sequence and said second binding partner moiety, wherein said first and second probes hybridize to target and reference nucleic acid sequences, respectively, present in said sample;
- (c) placing said hybridized probes under conditions that permit said binding partner moiety of said first probe to bind said binding partner moiety of said second probe, such that said first and second probes become bound to each other;
- and
- (d) detecting a probe that is not bound to another probe, wherein said detecting indicates a difference in the amount of target and reference nucleic acid sequences present in said sample, wherein said difference indicates the presence of a genetic abnormality.
A method of detecting a chromosomal abnormality, the method comprising:
- (a) contacting a nucleic acid sample with a probe under conditions that permit hybridization, wherein said probe comprises a sequence that specifically binds to said target nucleic acid sequence and a sequence that specifically binds to said reference nucleic acid sequence;
- (b) removing or rendering hybridized probes generated in step (a) resistant to detection;
- and
- (c) detecting unhybridized probe, target or reference nucleic acid, wherein said detecting determines the amount of said target nucleic acid relative to the amount of said reference nucleic acid present in said nucleic acid sample.
Dependencies are inferred from the claim text.