Method for detecting and amplifying target DNA
First Claim
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1. A method of detecting one or more nucleic acid target sequences in a sample, the method comprising the steps of:
- forming one or more closed circular probes whenever a first targeting domain and a second targeting domain of any of a plurality of precircle probes hybridize with their respective first target domains and second target domains of the one or more target sequences such that 5′ and
3′
nucleotides of the respective precircle probes abut one another, each precircle probe further having at least a first universal primer site and a cleavage site;
cleaving the one or more closed circular probes to form cleaved probes;
amplifying cleaved probes using a first universal primer complementary with a first universal primer site to form one or more amplicons; and
detecting the amplicons to detect the one or more nucleic acid target sequences in the sample.
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Abstract
The invention is directed to novel methods of multiplexing nucleic acid reactions, including amplification, detection and genotyping. The invention relies on the use of precircle probes that are circularized in the presence of the corresponding target nucleic acids, cleaved, and then amplified.
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Citations
24 Claims
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1. A method of detecting one or more nucleic acid target sequences in a sample, the method comprising the steps of:
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forming one or more closed circular probes whenever a first targeting domain and a second targeting domain of any of a plurality of precircle probes hybridize with their respective first target domains and second target domains of the one or more target sequences such that 5′ and
3′
nucleotides of the respective precircle probes abut one another, each precircle probe further having at least a first universal primer site and a cleavage site;cleaving the one or more closed circular probes to form cleaved probes; amplifying cleaved probes using a first universal primer complementary with a first universal primer site to form one or more amplicons; and detecting the amplicons to detect the one or more nucleic acid target sequences in the sample. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21)
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22. A method of amplifying one or more nucleic acid target sequences, the method comprising the steps of:
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forming one or more closed circular probes whenever a first targeting domain and a second targeting domain of any of a plurality of precircle probes hybridize with their respective first target domains and second target domains of the one or more target sequences such that 5′ and
3′
nucleotides of the respective precircle probes abut one another, each precircle probe further having at least a first universal primer site and a cleavage site;cleaving the one or more closed circular probes to form cleaved probes; amplifying cleaved probes using a first universal primer complementary with a first universal primer site to form one or more amplicons. - View Dependent Claims (23, 24)
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Specification