US5514546A
Expired / Lapsed
US
⏱️ 0.0 years remaining
Stem-loop oligonucleotides containing parallel and antiparallel binding domains
Assignee: University of Rochester
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Filing: 1993-09-01
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Granted: 1996-05-07
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Expires: 2013-09-01
Abstract
Abstract The present invention provides stem-loop oligonucleotides containing a double-stranded stem domain of at least about 2 base pairs and a single-stranded loop domain. The loop domains of the present oligonucleotides include at least one parallel binding (P) domain separated by at least about 3 nucleotides from a corresponding anti-parallel binding (AP) domain. Each P and corresponding AP domain of the present oligonucleotides can bind detectably to one strand of a defined nucleic acid target wherein the P domain binds in a parallel manner to the target and the corresponding AP domain binds in an anti-parallel manner to the target. The present stem-loop oligonucleotides can bind to both single-stranded and double-stranded target nucleic acids. The present invention also provides methods of using these oligonucleotides as well as kits and pharmaceutical compositions containing these oligonucleotides.
Inventors
- 👤 Eric T. Kool
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-11-05 22:37
CPC Classifications
C; C07; C07H; C07H21/00; C12; C12Q; C12Q1/00; C12Q1/68; C12Q1/6813; C12Q1/6816; C12Q1/6823; C12Q1/6839
Claims Analysis
Total Claims: 10
Claim 1
Independent Claim
182 words
A method of detecting a target nucleic acid which comprises: contacting a sample to be tested for containing said target nucleic acid with a stem-loop oligonucleotide for a time and under conditions sufficient to form an oligonucleotide-target complex, wherein said stem-loop oligonucleotide comprises a double-stranded stem domain of at least about two base pairs and a single-stranded loop domain comprising at least one parallel binding (P) domain separated from at least one anti-parallel binding (AP) domain by a spacer having a length equivalent to at least about three nucleotides, wherein one P domain and one AP domain can simultaneously and detectably bind to one strand of said target nucleic acid, wherein said P domain is capable of binding in a parallel manner to said target by a sufficient number of Hoogsteen bonds to achieve detectable binding, and wherein said AP domain is capable of binding in an antiparallel manner to said target by a sufficient number of Watson-Crick bonds to achieve detectable binding; and detecting said complex, wherein detection of said complex is indicative of the presence of said target nucleic acid.
Claim 2
Independent Claim
182 words
A method of detecting a target nucleic acid which comprises: contacting a sample to be tested for containing said target nucleic acid with a stem-loop oligonucleotide for a time and under conditions sufficient to form an oligonucleotide-target complex, wherein said stem-loop oligonucleotide comprises a double-stranded stem domain of at least about two base pairs and a single-stranded loop domain comprising at least one parallel binding (P) domain separated from at least one anti-parallel binding (AP) domain by a spacer having a length equivalent to at least about three nucleotides, wherein one P domain and one AP domain can simultaneously and detectably bind to one strand of said target nucleic acid, wherein said P domain is capable of binding in a parallel manner to said target by a sufficient number of Hoogsteen bonds to achieve detectable binding, and wherein said AP domain is capable of binding in an antiparallel manner to said target by a sufficient number of Watson-Crick bonds to achieve detectable binding; and detecting said complex, wherein detection of said complex is indicative of the presence of said target nucleic acid.
Claim 3
Dependent Claim
44 words
The method of claim 1 wherein said target nucleic acid comprises a double-stranded nucleic acid target, and wherein said conditions are sufficient to denature said double-stranded nucleic acid target by strand displacement and thereby permit binding of said oligonucleotide to form said oligonucleotide-target complex.
Claim 4
Dependent Claim
24 words
The method of claim 1 wherein said sample comprises a mixture of nucleic acids, a tissue section, a cell smear or a chromosomal squash.
Claim 5
Dependent Claim
22 words
The method of claim 2 wherein said time effective to denature said target ranges from about 1 minute to about 16 hours.
Claim 6
Dependent Claim
16 words
The method of claim 1 wherein said complex is detected by a fluorescence energy transfer assay.
Claim 7
Dependent Claim
16 words
The method of claim 1 wherein said spacer is a nucleotide spacer or a non-nucleotide spacer.
Claim 8
Dependent Claim
12 words
The method of claim 6 wherein said non-nucleotide spacer is polyethylene glycol.
Claim 9
Dependent Claim
17 words
The method of claim 7 wherein said polyethylene glycol is pentaethylene glycol, hexaethylene glycol or heptaethylene glycol.
Claim 10
Dependent Claim
13 words
The method of claim 1 wherein said oligonucleotide further comprises a reporter molecule.
References Cited (Backward Citations)
US4868103A
Cited
US5422251A
Cited
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