Format of Probes to Detect Nucleic Acid Differences
First Claim
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1. A method of detecting the presence or absence of a target nucleic acid sequence in a biological sample, the method comprising:
- a. contacting a detectably-labeled probe comprising an anchor nucleic acid domain and a reporter nucleic acid domain with the sample; and
b. detecting the presence or absence of binding of the probe to the target nucleic acid,wherein the anchor and reporter domains are linked by a non-nucleoside linker, and neither the anchor nor the reporter domain forms a stem loop in the absence of the target nucleic acid;
and wherein(i) the probe is not extendible by a polymerase; and
/or(ii) the linker is linked to the anchor domain within 2 nucleotides of the 3′
end of the anchor domain and the linker is linked to the reporter domain within 2 nucleotides of the 5′
end of the reporter domain, wherein the anchor domain is not linked to a detectable label; and
/or(iii) the anchor domain and the reporter domain each comprise a contiguous sequence of at least 6 nucleotides complementary to the same strand of the target nucleic acid.
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Abstract
The invention provides, inter alia, novel probes, methods, reaction mixtures, and kits for detecting the presence or absence of a target nucleic acid sequence.
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Citations
36 Claims
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1. A method of detecting the presence or absence of a target nucleic acid sequence in a biological sample, the method comprising:
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a. contacting a detectably-labeled probe comprising an anchor nucleic acid domain and a reporter nucleic acid domain with the sample; and b. detecting the presence or absence of binding of the probe to the target nucleic acid, wherein the anchor and reporter domains are linked by a non-nucleoside linker, and neither the anchor nor the reporter domain forms a stem loop in the absence of the target nucleic acid; and wherein (i) the probe is not extendible by a polymerase; and
/or(ii) the linker is linked to the anchor domain within 2 nucleotides of the 3′
end of the anchor domain and the linker is linked to the reporter domain within 2 nucleotides of the 5′
end of the reporter domain, wherein the anchor domain is not linked to a detectable label; and
/or(iii) the anchor domain and the reporter domain each comprise a contiguous sequence of at least 6 nucleotides complementary to the same strand of the target nucleic acid. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12)
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13. A reaction mixture for detecting the presence or absence of a target sequence comprising:
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a. a target nucleic acid comprising an anchor binding region and a reporter binding region, and b. a detectably-labeled probe comprising an anchor nucleic acid domain and a reporter nucleic acid domain, wherein the anchor and reporter domains are linked by a non-nucleoside linker, and neither the anchor nor the reporter domain forms a stem loop in the absence of the target nucleic acid, and wherein (i) the probe is not extendible by a polymerase; (ii) the linker is linked to the anchor domain within 2 nucleotides of the 3′
end of the anchor domain and the linker is linked to the reporter domain within 2 nucleotides of the 5′
end of the reporter domain, wherein the anchor domain is not linked to a detectable label; and
/or(iii) the anchor domain and the reporter domain each comprise a contiguous sequence of at least 6 nucleotides complementary to the same strand of the target nucleic acid. - View Dependent Claims (14, 15, 16, 17, 18, 19, 20, 21, 22)
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23. A detectably-labeled probe comprising an anchor nucleic acid domain and a reporter nucleic acid domain, wherein:
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the anchor and reporter domains are linked by a non-nucleoside linker; neither the anchor nor the reporter domain forms a stem loop in the absence of the target nucleic acid; and wherein; (i) the probe is not extendible by a polymerase; and
/or(ii) the linker is linked to the anchor domain within 2 nucleotides of the 3′
end of the anchor domain and the linker is linked to the reporter domain within 2 nucleotides of the 5′
end of the reporter domain, wherein the anchor domain is not linked to a detectable label. - View Dependent Claims (24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36)
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Specification