Methods and Compositions to Detect Nucleic Acids in a Biological Sample
First Claim
1. A kit that includes a capture probe having the structure 5′
- -X.sub.n a′
b′
c′
Y.sub.n-3′
, wherein X and Y each comprise nucleic acid sequences that can form a double stranded portion of a hairpin structure and wherein one of X or Y is a capture sequence further comprising a first member of a specific binding pair and the other of X or Y is a terminal sequence of the hairpin capture probe, and wherein a′
b′
c′
comprises a target-complementary sequence and is flanked by X and Y to thereby form a loop portion of the hairpin structure; and
a detection probe that hybridizes specifically to a target nucleic acid sequence that is at least partially the same as the sequence hybridized by the a′
b′
c′
portion of the capture probe.
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Accused Products
Abstract
Kits, reaction mixtures and methods for separating a target nucleic acid from a sample by using at least one hairpin capture probe oligonucleotide that has the structure 5′-X.sub.n a′ b′ c′ Y.sub.n-3′, wherein X and Y each comprise nucleic acid sequences that can form a double stranded stem portion, one of X or Y is a capture sequence that is a first member of a specific binding pair and the other of X or Y is a terminal sequence of the hairpin capture probe, and a′ b′ c′ comprises a target-complementary sequence flanked by X and Y to thereby form a loop portion of the hairpin, thus forming a capture hybrid that is separated from other sample components before the target nucleic acid is released from the capture support and hybridized to a detection probe that hybridizes specifically to the same sequence that is at least partially hybridized by the a′ b′ c′ portion of the capture probe, thus forming a detectable detection hybrid to indicate the presence of the target nucleic acid in the sample.
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Citations
24 Claims
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1. A kit that includes a capture probe having the structure 5′
- -X.sub.n a′
b′
c′
Y.sub.n-3′
, wherein X and Y each comprise nucleic acid sequences that can form a double stranded portion of a hairpin structure and wherein one of X or Y is a capture sequence further comprising a first member of a specific binding pair and the other of X or Y is a terminal sequence of the hairpin capture probe, and wherein a′
b′
c′
comprises a target-complementary sequence and is flanked by X and Y to thereby form a loop portion of the hairpin structure; and
a detection probe that hybridizes specifically to a target nucleic acid sequence that is at least partially the same as the sequence hybridized by the a′
b′
c′
portion of the capture probe. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10)
- -X.sub.n a′
-
11. A reaction mixture for the detection of a target nucleic acid, wherein the reaction mixture comprises:
- a capture probe having the structure 5′
-X.sub.n a′
b′
c′
Y.sub.n-3′
, wherein X and Y each comprise nucleic acid sequences that can form a double stranded portion of a hairpin structure and wherein one of X or Y is a capture sequence further comprising a first member of a specific binding pair and the other of X or Y is a terminal sequence of the hairpin capture probe, and wherein a′
b′
c′
comprises a target-complementary sequence and is flanked by X and Y to thereby form a loop portion of the hairpin structure; and
a detection probe that hybridizes specifically to a target nucleic acid sequence that is at least partially the same as the sequence hybridized by the a′
b′
c′
portion of the capture probe. - View Dependent Claims (12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23)
- a capture probe having the structure 5′
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24. A method for the capture and detection of a target nucleic acid, comprising the steps of:
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a. providing to a sample that is suspected of containing a target nucleic acid, a capture probe that has the structure 5′
-X.sub.n a′
b′
c′
Y.sub.n-3′
, wherein X and Y each comprise nucleic acid sequences that can form a double stranded portion of a hairpin structure, one of X or Y is a capture sequence further comprising a first member of a specific binding pair and the other of X or Y is a terminal sequence of the hairpin capture probe, and a′
b′
c′
comprises a target-complementary sequence that is flanked by X and Y to thereby form a loop portion of the hairpin structure;b. providing conditions for capture of the target nucleic acid, wherein the conditions are configured to hybridize the a′
b′
c′
portion of the capture probe to a sequence of the target nucleic acid in the sample and wherein the X and Y members are not hybridized one to the other;c. providing to a reaction mixture comprising the target nucleic acid and the capture probe a detection probe that is configured to hybridize specifically to the target nucleic acid at a sequence that is at least partially the same as the sequence that the a′
b′
c′
portion of the capture probe hybridizes;d. providing conditions configured to form a hairpin capture probe wherein X and Y form the double stranded stem and a′
b′
c′
is the single stranded loop, and wherein the detection probe is hybridized to the a sequence that is at least partially the same as the sequence that the a′
b′
c′
portion of the capture probe hybridizes; ande. detecting a signal from the detection probe to determine the presence or absence of the target nucleic acid from the sample.
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Specification