DETECTION OF NUCLEIC ACIDS BY TYPE SPECIFIC HYBRID CAPTURE METHOD
First Claim
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1. A method of detecting a target nucleic acid comprising:
- a) hybridizing a single-stranded target nucleic acid to a capture sequence probe and a signal sequence probe to form double-stranded hybrids between said probes and the target nucleic acid, wherein the capture sequence probe and the signal sequence probe are capable of hybridizing to non-overlapping regions within the target nucleic acid and not being capable of hybridizing to each other;
b) adding a blocker probe to the hybridization reaction, wherein said blocker probe hybridizes to excess non-hybridized capture sequence probes;
c) capturing the hybrid to form a bound hybrid; and
d) detecting the bound hybrid.
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Abstract
Target-specific hybrid capture (TSHC) provides a nucleic acid detection method that is not only rapid and sensitive, but is also highly specific and capable of discriminating highly homologous nucleic acid target sequences. The method produces DNA-RNA hybrids which can be detected by a variety of methods.
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Citations
55 Claims
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1. A method of detecting a target nucleic acid comprising:
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a) hybridizing a single-stranded target nucleic acid to a capture sequence probe and a signal sequence probe to form double-stranded hybrids between said probes and the target nucleic acid, wherein the capture sequence probe and the signal sequence probe are capable of hybridizing to non-overlapping regions within the target nucleic acid and not being capable of hybridizing to each other; b) adding a blocker probe to the hybridization reaction, wherein said blocker probe hybridizes to excess non-hybridized capture sequence probes; c) capturing the hybrid to form a bound hybrid; and d) detecting the bound hybrid. - View Dependent Claims (3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 32, 37, 38, 39, 48, 49)
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2. A method of detecting a target nucleic acid comprising:
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a) hybridizing a single-stranded target nucleic acid to an immobilized capture sequence probe and a signal sequence probe to form double-stranded hybrids between said probes and the target nucleic acid, wherein the capture sequence probe and the signal sequence probe are capable of hybridizing to non-overlapping regions within the target nucleic acid and not being capable of hybridizing to each other; b) adding a blocker probe to the hybridization reaction, wherein said blocker probe hybridizes to excess non-hybridized capture sequence probes; c) detecting the bound hybrid.
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22. A method of detecting a target nucleic acid comprising:
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a) hybridizing a single-stranded target nucleic acid to a capture sequence probe and a signal sequence probe, wherein the capture sequence probe and the signal sequence probe are capable of hybridizing to non-overlapping regions within the target nucleic acid and not being capable of hybridizing to each other, wherein said hybridization forms an RNA-DNA hybrid between said signal sequence probe and the target nucleic acid; and b) detecting the RNA-DNA hybrid by binding an antibody capable of recognizing the RNA-DNA hybrid to said hybrid, wherein said antibody is detectably labelled. - View Dependent Claims (23, 24, 25, 26, 27, 28, 29, 30, 31, 33, 34, 35, 36)
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40. A method of detecting a target nucleic acid comprising:
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a) hybridizing a single stranded target nucleic acid to a capture sequence probe and a signal sequence probe, wherein the capture sequence probe and the signal sequence probe are capable of hybridizing to non-overlapping regions within the target nucleic acid and not being capable of hybridizing to each other, wherein the signal sequence probe comprises a DNA-RNA hybrid region, wherein said hybridization forms a complex; and b) detecting said complex. - View Dependent Claims (41, 42, 43, 44, 45, 46)
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47. A nucleic acid probe consisting of a sequence selected from the group consisting of SEQ ID NO:
- 1 through SEQ ID NO;
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- 1 through SEQ ID NO;
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50. A method of detecting a target nucleic acid comprising:
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a) hybridizing a single-stranded target nucleic acid to a capture sequence probe, a bridge probe and a signal sequence probe to form double-stranded hybrids between said capture sequence and bridge probes and the target nucleic acid, wherein the capture sequence probe and the bridge probe are capable of hybridizing to non-overlapping regions within the target nucleic acid and not being capable of hybridizing to each other, and the signal sequence probe is capable of hybridizing to the bridge probe and not being capable of hybridizing to the target nucleic acid and the capture sequence probe; b) adding a blocker probe to the hybridization reaction, wherein said blocker probe hybridizes to excess non-hybridized capture sequence probes; c) capturing the hybrid to form a bound hybrid; and d) detecting the bound hybrid. - View Dependent Claims (51, 52, 53, 54, 55)
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Specification