EP0232967B1
Expired / Lapsed
EP
⏱️ 0.0 years remaining
Competitive homogeneous assay
Assignee: BP Corp North America Inc
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Filing: 1987-01-09
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Granted: 1993-04-28
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Expires: 2007-01-09
Abstract
Abstract A method for assaying a sample for target polynucleotides comprises (a) contacting the sample with a reagent under binding conditions where the reagent includes a first polynucleotide probe (I) and a second polynucleotide probe (II). (I) and (II) capable of assuming a first position where (I) and (II) are bound to each other and at least one of the probes capable of assuming a second position where the probe strand is bound to the target, (I) and (II) including a first label moiety (L1) associated with one of the probes and including a second label moiety (L2) associated with the opposite probe, L1 and L2 capable of interacting when (I) and (II) are bound to each other to produce a signal capable of detection characteristic of the probe in one of the 2 positions and (b) monitoring the sample for the signal, the presence of which is related to the presence of target in the sample. Also claimed is a method for associating an amine-reactive moiety to the 3'-terminus of a polynucleotide comprising (a) reacting the polynucleotide with an aminoalkyl deriv. of a nucleic acid in the presence of the enzyme terminal transferase, (b) reacting the amino gp. of the aminoalkyl deriv. with the amine-reactive moiety. A prefd. aminoalkyl deriv. is 8-(6-amino-hexyl)aminoadenosine -5'-triphosphate.
Inventors
- 👤 Larry Edward Morrison
Bibliographic Details
Application Number
—
Jurisdiction
EP
Data Source
google_patents
Last Synchronized
2026-11-05 05:52
CPC Classifications
C; C12; C12Q; C12Q1/00; C12Q1/68; C12Q1/6813; C12Q1/6816; C12Q1/6818; C07; C07H; C07H21/00; G; G01; G01N; G01N33/00; G01N33/48; G01N33/50; G01N33/58; G01N33/582
Claims Analysis
Total Claims: 25
Claim 1
Independent Claim
169 words
A method for assaying a sample for target polynucleotides comprising: (a) contacting the sample with reagent under binding conditions wherein said reagent includes a first polynucleotide probe and a second complementary polynucleotide probe, said first and second probes being capable of assuming a first position wherein said first and second probes are able to form a duplex and at least one of said probes being capable of assuming a second position wherein said at least one probe is bound to and forms a duplex with said target, said first and second probes including a first label moiety associated with one of said probes and including a second label moiety associated with said complementary probe, said first and second label moieties being capable of interacting, when said first and second probes form a duplex to produce a specific detectable signal characteristic of the probe in one of said two positions, and (b) monitoring said sample for said specific detectable signal, characteristic of the probe in one of said two positions.
Claim 2
Dependent Claim
35 words
The method of claim 1 wherein said first label moiety is located at the 3'-terminus of one of said probes and said second label moiety is located at the 5'-terminus of the said complementary probe.
Claim 3
Dependent Claim
17 words
The method of claim 1 or claim 2 wherein each probe has a plurality of label moieties.
Claim 4
Independent Claim
19 words
The method of any preceding claim wherein each such label moiety is located at the termini of said probes.
Claim 5
Independent Claim
24 words
The method of any preceding claim wherein said first label moiety is associated to said probe by an aminoalkyl derivative of a nucleic acid.
Claim 6
Dependent Claim
14 words
The method of claim 5 wherein said derivative includes an aminoalkyl derivative of adenine.
Claim 7
Dependent Claim
14 words
The method of claim 6 wherein said derivative includes an aminohexyl derivative of adenine.
Claim 8
Dependent Claim
10 words
The method of claim 6 wherein said derivative includes 8-(6-aminohexyl)-aminoadenosine-5'-monophosphate.
Claim 9
Independent Claim
25 words
The method of any of claims 1 to 7 wherein said label moiety at the 3'- terminus is a fluorescent derivative of a nucleic acid.
Claim 10
Dependent Claim
14 words
The method of claim 9 wherein the label moiety includes a derivative of 1-N⁶-ethenoadenosine-5'-monophosphate.
Claim 11
Independent Claim
50 words
A method for associating an amine-reactive moiety to the 3'-terminus of a polynucleotide comprising: (a) reacting said polynucleotide with an aminoalkyl derivative of a nucleic acid in the presence of the enzyme terminal transferase under reacting conditions. (b) reacting the amino group of the aminoalkyl derivative with said amine-reactive moiety.
Claim 12
Dependent Claim
15 words
The method of claim 11 wherein said aminoalkyl derivative includes an aminoalkyl derivative of adenine.
Claim 13
Dependent Claim
15 words
The method of claim 11 or claim 12 wherein said aminoalkyl derivative is a ribonucleotide.
Claim 14
Independent Claim
15 words
The method of any of claims 11 to 13 wherein said aminoalkyl derivative includes 8-(6-aminohexyl)-aminoadenosine-5'-triphosphate.
Claim 15
Independent Claim
172 words
A method for preparing polynucleotides including a first polynucleotide probe and a second complementary polynucleotide probe, said first and second probes being capable of assuming a first position wherein said first and second probes are able to form a duplex and at least one of said probes being capable of assuming a second position wherein said at least one probe is bound to and forms a duplex with a target, said first and second probes including a first label moiety associated with one of said probes and including a second label moiety associated with said complementary probe, said first and second label moieties being capable of interacting, when said first and second probes form a duplex to produce a specific detectable signal characteristic of the probe in one of said two positions, comprising: splicing polynucleotide segments having base sequences substantially identical to said target sequences into amplification means to form multiple copies of said polynucleotide segments, isolating said segments, and associating label moieties to the termini of the segments to form probes.
Claim 16
Dependent Claim
13 words
The method of claim 15 wherein amplification means include plasmids and phage particles.
Claim 17
Dependent Claim
34 words
The method of claim 15 or claim 16 wherein said segments, after isolation, are subjected to restriction digestion to form further subsegments, and said subsections are associated with said label moieties to form probes.
Claim 18
Independent Claim
49 words
The method of any of claims 15 to 17 wherein a said first label moiety is associated with the 3'-termini by reacting said segments with an aminoalkyl derivative of a nucleotide in the presence of terminal transferase under reacting conditions, and reacting said aminoalkyl group with a label moiety.
Claim 19
Dependent Claim
45 words
The method of claim 18 wherein a second label moiety is associated with the 5'-termini by reacting said segments with a bifunctional alkyl amine to form a segment with an amino alkyl group and thereafter reacting a second label moiety with said amino alkyl group.
Claim 20
Dependent Claim
142 words
A kit for assaying a sample for target polynucleotides comprising reagent, wherein said reagent includes a first polynucleotide probe and a second complementary polynucleotide probe, said first and second probes being capable of assuming a first position wherein said first and second probes are able to form a duplex and at least one of said probes being capable of assuming a second position wherein said at least one probe strand is bound to and forms a duplex with a target, said first and second probes including a first label moiety associated with one of said probes and including a second label moiety associated with said complementary probe, said first and second label moieties being capable of interacting when said first and second probes form a duplex to produce a specific detectable signal characteristic of the probe in one of said two positions.
Claim 21
Dependent Claim
25 words
The kit of claim 21 wherein at least one of said probes and/or label moieties is as defined in any of claims 2 to 9.
Claim 22
Independent Claim
39 words
A method according to any preceeding claim carried out in apparatus comprising: (a) chamber means adapted for receiving reagent and sample; (b) means for exciting one of said label moieties; and (c) means for detecting said specific detectable signal.
Claim 23
Dependent Claim
25 words
The method of claim 22 wherein said label moieties are fluorophores and said means for exciting one of said label moieties include a light source.
Claim 24
Dependent Claim
36 words
The method of claim 22 wherein at least one of said label moieties is a chemiluminescent agent and said means for exciting one of said label moieties include means for introducing chemiluminescent cofactors into said chamber.
Claim 25
Independent Claim
17 words
The method of any of claims 22 to 24 wherein said detection means include a light detector.
References Cited (Backward Citations)
No citation records indexed.
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