Detection of nucleic acids through amplification of surrogate nucleic acids
First Claim
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1. A method of detecting a first target nucleic acid, the method comprising:
- providing a sample comprising the first target nucleic acid;
associating the first target nucleic acid with a solid support byproviding a first set of m target capture probes, where m is at least one, which target capture probes are capable of hybridizing to the first target nucleic acid,hybridizing the first set of target capture probes with the first target nucleic acid, andassociating the first set of target capture probes with the solid support;
providing a first surrogate nucleic acid;
providing a first set of two or more surrogate capture probes, each of which surrogate capture probes is capable of hybridizing simultaneously to the first target nucleic acid and to a nucleic acid comprising the first surrogate nucleic acid;
hybridizing the first set of surrogate capture probes to the first target nucleic acid and to the nucleic acid comprising the first surrogate nucleic acid, thereby physically associating the first surrogate nucleic acid with the first target nucleic acid, to provide captured first surrogate nucleic acid;
amplifying the captured first surrogate nucleic acid, without amplifying the first target nucleic acid or a portion thereof, to provide amplified first surrogate nucleic acid; and
detecting the amplified first surrogate nucleic acid, whereby the presence or amount of the amplified first surrogate nucleic acid detected provides an indication of presence or amount of the first target nucleic acid in the sample;
wherein one or more copies of the nucleic acid comprising the first surrogate nucleic acid are associated with a copy of the first target nucleic acid;
wherein the first set of surrogate capture probes comprises a subset of surrogate capture probes for each of the one or more copies of the nucleic acid comprising the first surrogate nucleic acid;
wherein each subset of surrogate capture probes comprises n surrogate capture probes, where n is at least two; and
wherein the 5′
end of each of the surrogate capture probes in the first set hybridizes to the first target nucleic acid while the 3′
end hybridizes to the nucleic acid comprising the first surrogate nucleic acid, or wherein the 3′
end of each of the surrogate capture probes in the first set hybridizes to the first target nucleic acid while the 5′
end hybridizes to the nucleic acid comprising the first surrogate nucleic acid.
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Abstract
Methods for detecting and optionally quantitating one or more target nucleic acids are provided, in which a surrogate nucleic acid is captured to each target nucleic acid, amplified, and detected. Compositions, kit, and systems related to the methods are also described.
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22 Claims
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1. A method of detecting a first target nucleic acid, the method comprising:
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providing a sample comprising the first target nucleic acid; associating the first target nucleic acid with a solid support by providing a first set of m target capture probes, where m is at least one, which target capture probes are capable of hybridizing to the first target nucleic acid, hybridizing the first set of target capture probes with the first target nucleic acid, and associating the first set of target capture probes with the solid support; providing a first surrogate nucleic acid; providing a first set of two or more surrogate capture probes, each of which surrogate capture probes is capable of hybridizing simultaneously to the first target nucleic acid and to a nucleic acid comprising the first surrogate nucleic acid; hybridizing the first set of surrogate capture probes to the first target nucleic acid and to the nucleic acid comprising the first surrogate nucleic acid, thereby physically associating the first surrogate nucleic acid with the first target nucleic acid, to provide captured first surrogate nucleic acid; amplifying the captured first surrogate nucleic acid, without amplifying the first target nucleic acid or a portion thereof, to provide amplified first surrogate nucleic acid; and detecting the amplified first surrogate nucleic acid, whereby the presence or amount of the amplified first surrogate nucleic acid detected provides an indication of presence or amount of the first target nucleic acid in the sample; wherein one or more copies of the nucleic acid comprising the first surrogate nucleic acid are associated with a copy of the first target nucleic acid;
wherein the first set of surrogate capture probes comprises a subset of surrogate capture probes for each of the one or more copies of the nucleic acid comprising the first surrogate nucleic acid;
wherein each subset of surrogate capture probes comprises n surrogate capture probes, where n is at least two; and
wherein the 5′
end of each of the surrogate capture probes in the first set hybridizes to the first target nucleic acid while the 3′
end hybridizes to the nucleic acid comprising the first surrogate nucleic acid, or wherein the 3′
end of each of the surrogate capture probes in the first set hybridizes to the first target nucleic acid while the 5′
end hybridizes to the nucleic acid comprising the first surrogate nucleic acid. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22)
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Specification