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US6117635A
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Nucleic acid amplification oligonucleotides with molecular energy transfer labels and methods based thereon
Assignee: EMD Millipore Corp
•
Filing: 1997-04-11
•
Granted: 2000-09-12
•
Expires: 2017-04-11
Abstract
Abstract The present invention provides labeled nucleic acid amplification oligonucleotides, which can be linear or hairpin primers or blocking oligonucleotides. The oligonucleotides of the invention are labeled with donor and/or acceptor moieties of molecular energy transfer pairs. The moieties can be fluorophores, such that fluorescent energy emitted by the donor is absorbed by the acceptor. The acceptor may be a fluorophore that fluoresces at a wavelength different from the donor moiety, or it may be a quencher. The oligonucleotides of the invention are configured so that a donor moiety and an acceptor moiety are incorporated into the amplification product. The invention also provides methods and kits for directly detecting amplification products employing the nucleic acid amplification primers. When labeled linear primers are used, treatment with exonuclease or by using specific temperature eliminates the need for separation of unincorporated primers. This "closed-tube" format greatly reduces the possibility of carryover contamination with amplification products, provides for high throughput of samples, and may be totally automated.
Inventors
- 👤 Irina A. Nazarenko
- 👤 Satish K. Bhatnagar
- 👤 Emily S. Winn-Deen
- 👤 Robert J. Hohman
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-10-29 21:38
CPC Classifications
C; C12; C12Q; C12Q1/00; C12Q1/68; C12Q1/6813; C12Q1/6816; C12Q1/6818; C12Q1/6844; C12Q1/6858; C12Q1/686
Claims Analysis
Total Claims: 127
Claim 1
Independent Claim
35 words
An oligodeoxynucleotide, the sequence of which consists of: 5'-GGCTACGAACCAGGTAAGCCGTA-3' (SEQ ID NO:1), wherein fluorescein or a derivative thereof is attached to the 5' G and DABCYL is attached to the T at nucleotide number 22.
Claim 2
Dependent Claim
35 words
An oligodeoxynucleotide, the sequence of which consists of: 5'-GGCTACGAACCAGGTAAGCCGTA-3' (SEQ ID NO:1), wherein fluorescein or a derivative thereof is attached to the 5' G and DABCYL is attached to the T at nucleotide number 22.
Claim 3
Dependent Claim
35 words
An oligodeoxynucleotide, the sequence of which consists of: 5'-ACCTTCTGTTCACCCTCAGAAGGTGACCAAGTTCAT-3' (SEQ ID NO:14), wherein fluorescein or a derivative thereof is attached to the 5' A and DABCYL is attached to the T at nucleotide number 24.
Claim 4
Dependent Claim
35 words
An oligodeoxynucleotide, the sequence of which consists of: 5'-ACCTTCTGTTCACCCTCAGAAGGTGACCAAGTTCAT-3' (SEQ ID NO:14), wherein fluorescein or a derivative thereof is attached to the 5' A and DABCYL is attached to the T at nucleotide number 24.
Claim 5
Dependent Claim
35 words
An oligodeoxynucleotide, the sequence of which consists of: 5'-ACCTTCGATTCACCCTCAGAAGGTGACCAAGTTCAT-3' (SEQ ID NO:15), wherein fluorescein or a derivative thereof is attached to the 5' A and DABCYL is attached to the T at nucleotide number 24.
Claim 6
Dependent Claim
35 words
An oligodeoxynucleotide, the sequence of which consists of: 5'-ACCTTCGATTCACCCTCAGAAGGTGACCAAGTTCAT-3' (SEQ ID NO:15), wherein fluorescein or a derivative thereof is attached to the 5' A and DABCYL is attached to the T at nucleotide number 24.
Claim 7
Dependent Claim
35 words
An oligodeoxynucleotide, the sequence of which consists of: 5'-ACCTTCTGTACCCTCAGAAGGTGACCAAGTTCAT-3' (SEQ ID NO:16), wherein fluorescein or a derivative thereof is attached to the 5' A and DABCYL is attached to the T at nucleotide number 22.
Claim 8
Dependent Claim
35 words
An oligodeoxynucleotide, the sequence of which consists of: 5'-ACCTTCTGTACCCTCAGAAGGTGACCAAGTTCAT-3' (SEQ ID NO:16), wherein fluorescein or a derivative thereof is attached to the 5' A and DABCYL is attached to the T at nucleotide number 22.
Claim 9
Dependent Claim
35 words
An oligodeoxynucleotide, the sequence of which consists of: 5'-ACCTTCTATACCCTCAGAAGGTGACCAAGTTCAT-3' (SEQ ID NO:17), wherein fluorescein or a derivative thereof is attached to the 5' A and DABCYL is attached to the T at nucleotide number 22.
Claim 10
Dependent Claim
35 words
An oligodeoxynucleotide, the sequence of which consists of: 5'-ACCTTCTATACCCTCAGAAGGTGACCAAGTTCAT-3' (SEQ ID NO:17), wherein fluorescein or a derivative thereof is attached to the 5' A and DABCYL is attached to the T at nucleotide number 22.
Claim 11
Dependent Claim
139 words
An oligonucleotide containing: (a) a first nucleotide sequence, (b) a second nucleotide sequence at the 5' end of the first nucleotide sequence, (c) a third nucleotide sequence at the 5' end of the second nucleotide sequence, and (d) a fourth nucleotide sequence at the 5' end of the third nucleoide sequence, wherein the first nucleotide sequence contains the nucleotide at the 3' end of the oligonucleotide and the fourth nucleotide sequence contains the nucleotide at the 5' end of the oligonucleotide, wherein a plurality of nucleotides of the first nucleotide sequence is unable to form part of a hairpin capable of being formed by the oligonucleotide, wherein, if the hairpin is formed, then the hairpin contains nucleotides of the second and fourth nucleotide sequences, and wherein, if the hairpin is not formed, then the oligonucleotide emits a detectable signal.
Claim 12
Dependent Claim
139 words
An oligonucleotide containing: (a) a first nucleotide sequence, (b) a second nucleotide sequence at the 5' end of the first nucleotide sequence, (c) a third nucleotide sequence at the 5' end of the second nucleotide sequence, and (d) a fourth nucleotide sequence at the 5' end of the third nucleoide sequence, wherein the first nucleotide sequence contains the nucleotide at the 3' end of the oligonucleotide and the fourth nucleotide sequence contains the nucleotide at the 5' end of the oligonucleotide, wherein a plurality of nucleotides of the first nucleotide sequence is unable to form part of a hairpin capable of being formed by the oligonucleotide, wherein, if the hairpin is formed, then the hairpin contains nucleotides of the second and fourth nucleotide sequences, and wherein, if the hairpin is not formed, then the oligonucleotide emits a detectable signal.
Claim 13
Dependent Claim
18 words
The oligonucleotide of claim 6, wherein the first nucleotide sequence is not complementary to the second nucleotide sequence.
Claim 14
Dependent Claim
18 words
The oligonucleotide of claim 6, wherein the first nucleotide sequence is not complementary to the third nucleotide sequence.
Claim 15
Dependent Claim
34 words
The oligonucleotide of claim 6, wherein the detectable signal emitted by the oligonucleotide if the hairpin is not formed is more intense than a signal emitted by the oligonucleotide if the hairpin is formed.
Claim 16
Dependent Claim
19 words
The oligonucleotide of claim 6, wherein the oligonucleotide emits the detectable signal only if the hairpin is not formed.
Claim 17
Dependent Claim
11 words
The oligonucleotide of claim 10, wherein the detectable signal is substantial.
Claim 18
Dependent Claim
112 words
The oligonucleotide of claim 6 further containing a molecular energy transfer pair including an energy donor moiety that is capable of emitting energy, and an energy acceptor moiety that is capable of absorbing an amount of the emitted energy, wherein the donor moiety is attached to a nucleotide of the second nucleotide sequence and the acceptor moiety is attached to a nucleotide of the fourth nucleotide sequence, or the acceptor moiety is attached to a nucleotide of the second nucleotide sequence and the donor moiety is attached to a nucleotide of the fourth nucleotide sequence; and the acceptor moiety absorbs the amount of the emitted energy only if the hairpin is formed.
Claim 19
Dependent Claim
21 words
The oligonucleotide of claim 6, wherein the first nucleotide sequence is complementary to a nucleotide sequence flanking a target nucleotide sequence.
Claim 20
Dependent Claim
13 words
The oligonucleotide of claim 13, wherein the target nucleotide sequence is genomic DNA.
Claim 21
Dependent Claim
12 words
The oligonucleotide of claim 13, wherein the target nucleotide sequence is cDNA.
Claim 22
Dependent Claim
12 words
The oligonucleotide of claim 13, wherein the target nucleotide sequence is mRNA.
Claim 23
Dependent Claim
14 words
The oligonucleotide of claim 13, wherein the target nucleotide sequence is chemically synthesized DNA.
Claim 24
Dependent Claim
18 words
The oligonucleotide of claim 13, wherein the target nucleotide sequence is a sequence of an infectious disease agent.
Claim 25
Dependent Claim
30 words
The oligonucleotide of claim 13, wherein the target nucleotide sequence is a wild-type human genomic sequence, mutation of which is implicated in the presence of a human disease or disorder.
Claim 26
Dependent Claim
14 words
The method of claim 13, wherein the target nucleotide sequence is an amplification product.
Claim 27
Dependent Claim
15 words
The method of claim 20, wherein the amplification product contains a restriction endonuclease recognition site.
Claim 28
Dependent Claim
25 words
The oligonucleotide of claim 12, wherein the donor moiety is a fluorophore and the acceptor moiety is a quencher of light emitted by the fluorophore.
Claim 29
Dependent Claim
11 words
The oligonucleotide of claim 12, wherein the donor moiety is fluorescein.
Claim 30
Dependent Claim
14 words
The oligonucleotide of claim 12, wherein the donor moiety is a derivative of fluorescein.
Claim 31
Dependent Claim
11 words
The oligonucleotide of claim 12, wherein the donor moiety is 5-carboxyfluorescein.
Claim 32
Dependent Claim
11 words
The oligonucleotide of claim 12, wherein the donor moiety is rhodamine.
Claim 33
Dependent Claim
13 words
The oligonucleotide of claim 12, wherein the donor moiety is 5-(2'-aminoethyl) aminonapthalene-1-sulfonic acid.
Claim 34
Dependent Claim
11 words
The oligonucleotide of claim 12, wherein the donor moiety is anthranilamide.
Claim 35
Dependent Claim
11 words
The oligonucleotide of claim 12, wherein the donor moiety is coumarin.
Claim 36
Dependent Claim
13 words
The oligonucleotide of claim 12, wherein the donor moiety is a metal chelate.
Claim 37
Dependent Claim
14 words
The oligonucleotide of claim 12, wherein the donor moiety is a terbium chelate derivative.
Claim 38
Dependent Claim
13 words
The oligonucleotide of claim 12, wherein the donor moiety is Reactive Red 4.
Claim 39
Dependent Claim
11 words
The oligonucleotide of claim 12, wherein the acceptor moiety is DABCYL.
Claim 40
Dependent Claim
11 words
The oligonucleotide of claim 12, wherein the acceptor moiety is rhodamine.
Claim 41
Dependent Claim
12 words
The oligonucleotide of claim 12, wherein the acceptor moiety is pyrene butyrate.
Claim 42
Dependent Claim
12 words
The oligonucleotide of claim 12, wherein the acceptor moiety is eosine nitrotyrosine.
Claim 43
Dependent Claim
11 words
The oligonucleotide of claim 12, wherein the acceptor moiety is ethidium.
Claim 44
Dependent Claim
11 words
The oligonucleotide of claim 12, wherein the acceptor moiety is fluorescein.
Claim 45
Dependent Claim
14 words
The oligonucleotide of claim 12, wherein the acceptor moiety is a derivative of fluorescein.
Claim 46
Dependent Claim
12 words
The oligonucleotide of claim 12, wherein the acceptor moiety is Malachite green.
Claim 47
Dependent Claim
12 words
The oligonucleotide of claim 12, wherein the acceptor moiety is Texas Red.
Claim 48
Dependent Claim
17 words
The oligonucleotide of claim 12, wherein the donor moiety is fluorescein and the acceptor moiety is DABCYL.
Claim 49
Dependent Claim
20 words
The oligonucleotide of claim 12, wherein the donor moiety is a derivative of fluorescein and the acceptor moiety is DABCYL.
Claim 50
Dependent Claim
27 words
The oligonucleotide of claim 12, wherein the nucleotide to which the donor moiety is attached is complementary to the nucleotide to which the acceptor moiety is attached.
Claim 51
Dependent Claim
38 words
The oligonucleotide of claim 12, wherein, if the hairpin is formed, then the nucleotide to which the donor moiety is attached and the complement of the nucleotide to which the acceptor moiety is attached are five nucleotides apart.
Claim 52
Dependent Claim
32 words
The oligonucleotide of claim 12, wherein the nucleotide to which the donor moiety is attached and the nucleotide to which the acceptor moiety is attached are from 15 to 25 nucleotides apart.
Claim 53
Dependent Claim
11 words
The oligonucleotide of claim 6, wherein the oligonucleotide is an oligodeoxynucleotide.
Claim 54
Dependent Claim
16 words
The oligonucleotide of claim 6, wherein the first nucleotide sequence contains a restriction endonuclease recognition site.
Claim 55
Dependent Claim
10 words
The oligonucleotide of claim 6, wherein the signal is light.
Claim 56
Dependent Claim
38 words
An oligonucleotide comprising the nucleotide sequence of SEQ ID NO:1, wherein fluorescein is attached to the nucleotide at position 1 of SEQ ID NO:1, and DABCYL is attached to the nucleotide at position 22 of SEQ ID NO:1.
Claim 57
Dependent Claim
38 words
An oligonucleotide comprising the nucleotide sequence of SEQ ID NO:1, wherein fluorescein is attached to the nucleotide at position 1 of SEQ ID NO:1, and DABCYL is attached to the nucleotide at position 22 of SEQ ID NO:1.
Claim 58
Dependent Claim
14 words
The oligonucleotide of claim 50 consisting of the nucleotide sequence of SEQ ID NO:1.
Claim 59
Dependent Claim
41 words
An oligonucleotide comprising the nucleotide sequence of SEQ ID NO:1, wherein a derivative of fluorescein is attached to the nucleotide at position 1 of SEQ ID NO:1, and DABCYL is attached to the nucleotide at position 22 of SEQ ID NO:1.
Claim 60
Dependent Claim
41 words
An oligonucleotide comprising the nucleotide sequence of SEQ ID NO:1, wherein a derivative of fluorescein is attached to the nucleotide at position 1 of SEQ ID NO:1, and DABCYL is attached to the nucleotide at position 22 of SEQ ID NO:1.
Claim 61
Dependent Claim
14 words
The oligonucleotide of claim 6 consisting of the nucleotide sequence of SEQ ID NO:1.
Claim 62
Dependent Claim
38 words
An oligonucleotide comprising the nucleotide sequence of SEQ ID NO:13, wherein fluorescein is attached to the nucleotide at position 1 of SEQ ID NO:13, and DABCYL is attached to the nucleotide at position 20 of SEQ ID NO:13.
Claim 63
Dependent Claim
38 words
An oligonucleotide comprising the nucleotide sequence of SEQ ID NO:13, wherein fluorescein is attached to the nucleotide at position 1 of SEQ ID NO:13, and DABCYL is attached to the nucleotide at position 20 of SEQ ID NO:13.
Claim 64
Dependent Claim
14 words
The oligonucleotide of claim 54 consisting of the nucleotide sequence of SEQ ID NO:13.
Claim 65
Dependent Claim
41 words
An oligonucleotide comprising the nucleotide sequence of SEQ ID NO:13, wherein a derivative of fluorescein is attached to the nucleotide at position 1 of SEQ ID NO:13, and DABCYL is attached to the nucleotide at position 20 of SEQ ID NO:13.
Claim 66
Dependent Claim
41 words
An oligonucleotide comprising the nucleotide sequence of SEQ ID NO:13, wherein a derivative of fluorescein is attached to the nucleotide at position 1 of SEQ ID NO:13, and DABCYL is attached to the nucleotide at position 20 of SEQ ID NO:13.
Claim 67
Dependent Claim
14 words
The oligonucleotide of claim 56 consisting of the nucleotide sequence of SEQ ID NO:13.
Claim 68
Dependent Claim
38 words
An oligonucleotide comprising the nucleotide sequence of SEQ ID NO:14, wherein fluorescein is attached to the nucleotide at position 1 of SEQ ID NO:14, and DABCYL is attached to the nucleotide at position 24 of SEQ ID NO:14.
Claim 69
Dependent Claim
38 words
An oligonucleotide comprising the nucleotide sequence of SEQ ID NO:14, wherein fluorescein is attached to the nucleotide at position 1 of SEQ ID NO:14, and DABCYL is attached to the nucleotide at position 24 of SEQ ID NO:14.
Claim 70
Dependent Claim
14 words
The oligonucleotide of claim 58 consisting of the nucleotide sequence of SEQ ID NO:14.
Claim 71
Dependent Claim
41 words
An oligonucleotide comprising the nucleotide sequence of SEQ ID NO:14, wherein a derivative of fluorescein is attached to the nucleotide at position 1 of SEQ ID NO:14, and DABCYL is attached to the nucleotide at position 24 of SEQ ID NO:14.
Claim 72
Dependent Claim
41 words
An oligonucleotide comprising the nucleotide sequence of SEQ ID NO:14, wherein a derivative of fluorescein is attached to the nucleotide at position 1 of SEQ ID NO:14, and DABCYL is attached to the nucleotide at position 24 of SEQ ID NO:14.
Claim 73
Dependent Claim
14 words
The oligonucleotide of claim 60 consisting of the nucleotide sequence of SEQ ID NO:14.
Claim 74
Dependent Claim
38 words
An oligonucleotide comprising the nucleotide sequence of SEQ ID NO:15, wherein fluorescein is attached to the nucleotide at position 1 of SEQ ID NO:15, and DABCYL is attached to the nucleotide at position 24 of SEQ ID NO:15.
Claim 75
Dependent Claim
38 words
An oligonucleotide comprising the nucleotide sequence of SEQ ID NO:15, wherein fluorescein is attached to the nucleotide at position 1 of SEQ ID NO:15, and DABCYL is attached to the nucleotide at position 24 of SEQ ID NO:15.
Claim 76
Dependent Claim
14 words
The oligonucleotide of claim 62 consisting of the nucleotide sequence of SEQ ID NO:15.
Claim 77
Dependent Claim
41 words
An oligonucleotide comprising the nucleotide sequence of SEQ ID NO:15, wherein a derivative of fluorescein is attached to the nucleotide at position 1 of SEQ ID NO:15, and DABCYL is attached to the nucleotide at position 24 of SEQ ID NO:15.
Claim 78
Dependent Claim
41 words
An oligonucleotide comprising the nucleotide sequence of SEQ ID NO:15, wherein a derivative of fluorescein is attached to the nucleotide at position 1 of SEQ ID NO:15, and DABCYL is attached to the nucleotide at position 24 of SEQ ID NO:15.
Claim 79
Dependent Claim
14 words
The oligonucleotide of claim 64 consisting of the nucleotide sequence of SEQ ID NO:15.
Claim 80
Dependent Claim
38 words
An oligonucleotide comprising the nucleotide sequence of SEQ ID NO:16, wherein fluorescein is attached to the nucleotide at position 1 of SEQ ID NO:16, and DABCYL is attached to the nucleotide at position 22 of SEQ ID NO:16.
Claim 81
Dependent Claim
38 words
An oligonucleotide comprising the nucleotide sequence of SEQ ID NO:16, wherein fluorescein is attached to the nucleotide at position 1 of SEQ ID NO:16, and DABCYL is attached to the nucleotide at position 22 of SEQ ID NO:16.
Claim 82
Dependent Claim
14 words
The oligonucleotide of claim 66 consisting of the nucleotide sequence of SEQ ID NO:16.
Claim 83
Dependent Claim
41 words
An oligonucleotide comprising the nucleotide sequence of SEQ ID NO:16, wherein a derivative of fluorescein is attached to the nucleotide at position 1 of SEQ ID NO:16, and DABCYL is attached to the nucleotide at position 22 of SEQ ID NO:16.
Claim 84
Dependent Claim
41 words
An oligonucleotide comprising the nucleotide sequence of SEQ ID NO:16, wherein a derivative of fluorescein is attached to the nucleotide at position 1 of SEQ ID NO:16, and DABCYL is attached to the nucleotide at position 22 of SEQ ID NO:16.
Claim 85
Dependent Claim
14 words
The oligonucleotide of claim 68 consisting of the nucleotide sequence of SEQ ID NO:16.
Claim 86
Dependent Claim
38 words
An oligonucleotide comprising the nucleotide sequence of SEQ ID NO:17, wherein fluorescein is attached to the nucleotide at position 1 of SEQ ID NO:17, and DABCYL is attached to the nucleotide at position 22 of SEQ ID NO:17.
Claim 87
Dependent Claim
38 words
An oligonucleotide comprising the nucleotide sequence of SEQ ID NO:17, wherein fluorescein is attached to the nucleotide at position 1 of SEQ ID NO:17, and DABCYL is attached to the nucleotide at position 22 of SEQ ID NO:17.
Claim 88
Dependent Claim
14 words
The oligonucleotide of claim 70 consisting of the nucleotide sequence of SEQ ID NO:17.
Claim 89
Dependent Claim
41 words
An oligonucleotide comprising the nucleotide sequence of SEQ ID NO:17, wherein a derivative of fluorescein is attached to the nucleotide at position 1 of SEQ ID NO:17, and DABCYL is attached to the nucleotide at position 22 of SEQ ID NO:17.
Claim 90
Dependent Claim
41 words
An oligonucleotide comprising the nucleotide sequence of SEQ ID NO:17, wherein a derivative of fluorescein is attached to the nucleotide at position 1 of SEQ ID NO:17, and DABCYL is attached to the nucleotide at position 22 of SEQ ID NO:17.
Claim 91
Dependent Claim
14 words
The oligonucleotide of claim 72 consisting of the nucleotide sequence of SEQ ID NO:17.
Claim 92
Dependent Claim
38 words
An oligonucleotide comprising the nucleotide sequence of SEQ ID NO:18, wherein fluorescein is attached to the nucleotide at position 1 of SEQ ID NO:18, and DABCYL is attached to the nucleotide at position 20 of SEQ ID NO:18.
Claim 93
Dependent Claim
38 words
An oligonucleotide comprising the nucleotide sequence of SEQ ID NO:18, wherein fluorescein is attached to the nucleotide at position 1 of SEQ ID NO:18, and DABCYL is attached to the nucleotide at position 20 of SEQ ID NO:18.
Claim 94
Dependent Claim
14 words
The oligonucleotide of claim 74 consisting of the nucleotide sequence of SEQ ID NO:18.
Claim 95
Dependent Claim
41 words
An oligonucleotide comprising the nucleotide sequence of SEQ ID NO:18, wherein a derivative of fluorescein is attached to the nucleotide at position 1 of SEQ ID NO:18, and DABCYL is attached to the nucleotide at position 20 of SEQ ID NO:18.
Claim 96
Dependent Claim
41 words
An oligonucleotide comprising the nucleotide sequence of SEQ ID NO:18, wherein a derivative of fluorescein is attached to the nucleotide at position 1 of SEQ ID NO:18, and DABCYL is attached to the nucleotide at position 20 of SEQ ID NO:18.
Claim 97
Dependent Claim
14 words
The oligonucleotide of claim 76 consisting of the nucleotide sequence of SEQ ID NO:18.
Claim 98
Dependent Claim
90 words
A kit comprising: (a) a polymerase, and (b) an oligonucleotide containing: (i) a first nucleotide sequence, (ii) a second nucleotide sequence at the 5' end of the first nucleotide sequence, (iii) a third nucleotide sequence at the 5' end of the second nucleotide sequence, and (iv) a fourth nucleotide sequence at the 5' end of the third nucleotide sequence, wherein the oligonucleotide is capable of forming a hairpin containing nucleotides of the second and fourth nucleotide sequences, and the oligonucleotide emits a detectable signal if the hairpin is not formed.
Claim 99
Dependent Claim
90 words
A kit comprising: (a) a polymerase, and (b) an oligonucleotide containing: (i) a first nucleotide sequence, (ii) a second nucleotide sequence at the 5' end of the first nucleotide sequence, (iii) a third nucleotide sequence at the 5' end of the second nucleotide sequence, and (iv) a fourth nucleotide sequence at the 5' end of the third nucleotide sequence, wherein the oligonucleotide is capable of forming a hairpin containing nucleotides of the second and fourth nucleotide sequences, and the oligonucleotide emits a detectable signal if the hairpin is not formed.
Claim 100
Dependent Claim
47 words
The kit of claim 78, wherein the first nucleotide sequence contains the nucleotide at the 3' end of the oligonucleotide, the fourth nucleotide sequence contains the nucleotide at the 5' end of the oligonucleotide, and the first nucleotide sequence is not complementary to the fourth nucleotide sequence.
Claim 101
Dependent Claim
18 words
The kit of claim 79, wherein the first nucleotide sequence is not complementary to the second nucleotide sequence.
Claim 102
Dependent Claim
18 words
The kit of claim 79, wherein the first nucleotide sequence is not complementary to the third nucleotide sequence.
Claim 103
Dependent Claim
34 words
The kit of claim 78, wherein the detectable signal emitted by the oligonucleotide if the hairpin is not formed is more intense than a signal emitted by the oligonucleotide if the hairpin is formed.
Claim 104
Dependent Claim
19 words
The kit of claim 78, wherein the oligonucleotide emits the detectable signal only if the hairpin is not formed.
Claim 105
Dependent Claim
11 words
The kit of claim 83, wherein the detectable signal is substantial.
Claim 106
Dependent Claim
115 words
The kit of claim 78, wherein the oligonucleotide further contains a molecular energy transfer pair including an energy donor moiety that is capable of emitting energy, and an energy acceptor moiety that is capable of absorbing an amount of the emitted energy, wherein the donor moiety is attached to a nucleotide of the second nucleotide sequence and the acceptor moiety is attached to a nucleotide of the fourth nucleotide sequence, or the acceptor moiety is attached to a nucleotide of the second nucleotide sequence and the donor moiety is attached to a nucleotide of the fourth nucleotide sequence; and the acceptor moiety absorbs the amount of the emitted energy only if the hairpin is formed.
Claim 107
Dependent Claim
12 words
The kit of claim 78, wherein the polymerase is a DNA polymerase.
Claim 108
Dependent Claim
11 words
The kit of claim 78, wherein the oligonucleotide is an oligodeoxynucleotide.
Claim 109
Dependent Claim
21 words
The kit of claim 78 further comprising a ligase in a container that does not contain the polymerase or the oligonucleotide.
Claim 110
Dependent Claim
129 words
A kit comprising first and second oligonucleotides, wherein the first oligonucleotide contains: (i) a first nucleotide sequence complementary to a nucleotide sequence flanking a target nucleotide sequence, and (ii) a second nucleotide sequence at the 5" end of the first nucleotide sequence, and wherein the second oligonucleotide contains: (i) a third nucleotide sequence, (ii) a fourth nucleotide sequence at the 5' end of the third nucleotide sequence, (iii) a fifth nucleotide sequence at the 5' end of the fourth nucleotide sequence, and (iv) a sixth nucleotide sequence at the 5' end of the fifth nucleotide sequence, wherein the second oligonucleotide is capable of forming a hairpin containing nucleotides of the fourth and sixth nucleotide sequences, and the second oligonucleotide emits a detectable signal if the hairpin is not formed.
Claim 111
Dependent Claim
129 words
A kit comprising first and second oligonucleotides, wherein the first oligonucleotide contains: (i) a first nucleotide sequence complementary to a nucleotide sequence flanking a target nucleotide sequence, and (ii) a second nucleotide sequence at the 5" end of the first nucleotide sequence, and wherein the second oligonucleotide contains: (i) a third nucleotide sequence, (ii) a fourth nucleotide sequence at the 5' end of the third nucleotide sequence, (iii) a fifth nucleotide sequence at the 5' end of the fourth nucleotide sequence, and (iv) a sixth nucleotide sequence at the 5' end of the fifth nucleotide sequence, wherein the second oligonucleotide is capable of forming a hairpin containing nucleotides of the fourth and sixth nucleotide sequences, and the second oligonucleotide emits a detectable signal if the hairpin is not formed.
Claim 112
Dependent Claim
47 words
The kit of claim 89, wherein the third nucleotide sequence contains the nucleotide at the 3' end of the oligonucleotide, the sixth nucleotide sequence contains the nucleotide at the 5' end of the oligonucleotide, and the third nucleotide sequence is not complementary to the sixth nucleotide sequence.
Claim 113
Dependent Claim
18 words
The kit of claim 90, wherein the third nucleotide sequence is not complementary to the fourth nucleotide sequence.
Claim 114
Dependent Claim
18 words
The kit of claim 90, wherein the third nucleotide sequence is not complementary to the fifth nucleotide sequence.
Claim 115
Dependent Claim
116 words
The kit of claim 89, wherein the second oligonucleotide further contains a molecular energy transfer pair including an energy donor moiety that is capable of emitting energy, and an energy acceptor moiety that is capable of absorbing an amount of the emitted energy, wherein the donor moiety is attached to a nucleotide of the fourth nucleotide sequence and the acceptor moiety is attached to a nucleotide of the sixth nucleotide sequence, or the acceptor moiety is attached to a nucleotide of the fourth nucleotide sequence and the donor moiety is attached to a nucleotide of the sixth nucleotide sequence; and the acceptor moiety absorbs the amount of the emitted energy only if the hairpin is formed.
Claim 116
Dependent Claim
9 words
The kit of claim 89 further comprising a polymerase.
Claim 117
Dependent Claim
12 words
The kit of claim 94, wherein the polymerase is a DNA polymerase.
Claim 118
Dependent Claim
12 words
The kit of claim 89, wherein the first oligonucleotide is an oligodeoxynucleotide.
Claim 119
Dependent Claim
12 words
The kit of claim 89, wherein the second oligonucleotide is an oligodeoxynucleotide.
Claim 120
Dependent Claim
49 words
A kit comprising: (a) a polymerase, (b) a first oligonucleotide, and (c) a second oligonucleotide, wherein the first oligonucleotide is capable of annealing to the second oligonucleotide, and the first or second oligonucleotide emits a detectable signal only if the first oligonucleotide is not annealed to the second oligonucleotide.
Claim 121
Dependent Claim
49 words
A kit comprising: (a) a polymerase, (b) a first oligonucleotide, and (c) a second oligonucleotide, wherein the first oligonucleotide is capable of annealing to the second oligonucleotide, and the first or second oligonucleotide emits a detectable signal only if the first oligonucleotide is not annealed to the second oligonucleotide.
Claim 122
Dependent Claim
50 words
The kit of claim 98, wherein the detectable signal emitted by the first or second oligonucleotide if the first oligonucleotide is not annealed to the second oligonucleotide is more intense than a signal emitted by the first or second oligonucleotide if the first oligonucleotide is annealed to the second oligonucleotide.
Claim 123
Dependent Claim
27 words
The kit of claim 98, wherein the first or second oligonucleotide emits the detectable signal only if the first oligonucleotide is not annealed to the second oligonucleotide.
Claim 124
Dependent Claim
11 words
The kit of claim 100, wherein the detectable signal is substantial.
Claim 125
Dependent Claim
110 words
The kit of claim 98 further comprising a molecular energy pair including an energy donor moiety that is capable of emitting energy, and an energy acceptor moiety that is capable of absorbing an amount of the emitted energy, wherein the donor moiety is attached to a nucleotide of the first oligonucleotide and the acceptor moiety is attached to a nucleotide of the second oligonucleotide, or the acceptor moiety is attached to a nucleotide of the first oligonucleotide and the donor moiety is attached to a nucleotide of the second oligonucleotide, and the acceptor moiety absorbs the amount of energy only if the first oligonucleotide is annealed to the second oligonucleotide.
Claim 126
Dependent Claim
18 words
The oligonucleotide of claim 6, wherein the first nucleotide sequence is not complementary to the fourth nucleotide sequence.
Claim 127
Dependent Claim
15 words
The oligonucleotide of claim 6, wherein the oligonucleotide is capable of forming only one hairpin.
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Cited
USPTO Annuity & Maintenance Windows
| Annuity Stage | Window Opens | Payment Deadline | Surcharge Period Until | Estimated Fee | Status |
|---|---|---|---|---|---|
| 3.5 Years (Year 4) | 2003-09-12 | 2004-03-13 | 2004-09-12 | $2000.00 | Window Closed / Lapsed |
| 7.5 Years (Year 8) | 2007-09-12 | 2008-03-13 | 2008-09-12 | $3760.00 | Window Closed / Lapsed |
| 11.5 Years (Year 12) | 2011-09-12 | 2012-03-13 | 2012-09-12 | $7700.00 | Window Closed / Lapsed |