US6180348B1
Expired / Lapsed
US
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Method of isolating target specific oligonucleotide ligands
Assignee: BIOAGE PHARMACEUTICALS Inc
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Filing: 1999-04-07
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Granted: 2001-01-30
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Expires: 2019-04-07
Abstract
Abstract This invention relates to methods of preparing oligonucleotide libraries, isolating oligonucleotide aptamers to target molecules from the libraries, and using the aptamers to purify target molecules by affinity separation. Specific oligonucleotide aptamers to erythropoietin were isolated accordingly.
Inventors
- 👤 Weihua Li
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-10-26 02:06
CPC Classifications
C; C12; C12Q; C12Q1/00; C12Q1/68; C12Q1/6811
Claims Analysis
Total Claims: 16
Claim 1
Independent Claim
129 words
A method for identifying an aptamer which specifically binds to a target molecule other than a nucleic acid, comprising the steps of: a) providing a collection of oligonucleotides attached to a plurality of solid supports, wherein oligonucleotides of different nucleotide sequences are attached to different solid supports and all oligonucleotides attached to a single solid support are of the same nucleotide sequence; b) providing a plurality of magnetic beads having said target molecule immobilized on their surface; c) admixing said collection of oligonucleotides with said plurality of magnetic beads under conditions wherein the oligonucleotide ligands become bound to the target molecule; d) applying magnetic force to isolate one or more solid supports bound to magnetic beads; and e) identifying oligonucleotides attached to said one or more isolated solid supports.
Claim 2
Independent Claim
129 words
A method for identifying an aptamer which specifically binds to a target molecule other than a nucleic acid, comprising the steps of: a) providing a collection of oligonucleotides attached to a plurality of solid supports, wherein oligonucleotides of different nucleotide sequences are attached to different solid supports and all oligonucleotides attached to a single solid support are of the same nucleotide sequence; b) providing a plurality of magnetic beads having said target molecule immobilized on their surface; c) admixing said collection of oligonucleotides with said plurality of magnetic beads under conditions wherein the oligonucleotide ligands become bound to the target molecule; d) applying magnetic force to isolate one or more solid supports bound to magnetic beads; and e) identifying oligonucleotides attached to said one or more isolated solid supports.
Claim 3
Dependent Claim
25 words
The method of claim 1 , wherein said solid supports are resin beads and said step d) comprises selecting resin beads bound to magnetic beads.
Claim 4
Dependent Claim
21 words
The method of claim 2 , wherein said resin beads have a diameter of about 50 um to about 500 um.
Claim 5
Dependent Claim
21 words
The method of claim 2 , wherein said resin beads have a diameter of about 70 um to about 150 um.
Claim 6
Dependent Claim
21 words
The method of claim 2 , wherein said resin beads have a diameter of about 90 um to about 120 um.
Claim 7
Dependent Claim
21 words
The method of claim 1 , wherein said magnetic beads have a diameter of about 1 um to about 10 um.
Claim 8
Dependent Claim
21 words
The method of claim 1 , wherein said magnetic beads have a diameter of about 2 um to about 5 um.
Claim 9
Dependent Claim
16 words
The method of claim 1 , wherein said target molecule is labeled with a fluorescent probe.
Claim 10
Dependent Claim
19 words
The method of claim 1 , wherein said collection of oligonucleotides are synthesized on said plurality of solid supports.
Claim 11
Independent Claim
130 words
A method for identifying an aptamer which specifically binds to a target molecule other than a nucleic acid, comprising the steps of: a) providing a collection of oligonucleotides attached to a plurality of solid supports, wherein oligonucleotides of different nucleotide sequences are attached to different solid supports and all oligonucleotides attached to a single solid support are of the same nucleotide sequence; b) providing a plurality of said target molecules labeled with fluorescent probe and linked to a plurality of solid supports; c) admixing said collection of oligonucleotides with said plurality of said target molecules under conditions wherein the oligonucleotide ligands become bound to target molecule; d) isolating one or more solid supports attached with fluorescent probe; and e) identifying oligonucleotides attached to said one or more isolated solid supports.
Claim 12
Independent Claim
130 words
A method for identifying an aptamer which specifically binds to a target molecule other than a nucleic acid, comprising the steps of: a) providing a collection of oligonucleotides attached to a plurality of solid supports, wherein oligonucleotides of different nucleotide sequences are attached to different solid supports and all oligonucleotides attached to a single solid support are of the same nucleotide sequence; b) providing a plurality of said target molecules labeled with fluorescent probe and linked to a plurality of solid supports; c) admixing said collection of oligonucleotides with said plurality of said target molecules under conditions wherein the oligonucleotide ligands become bound to target molecule; d) isolating one or more solid supports attached with fluorescent probe; and e) identifying oligonucleotides attached to said one or more isolated solid supports.
Claim 13
Dependent Claim
17 words
The method of claim 10 , wherein said fluorescence labeled target molecules are linked to magetic beads.
Claim 14
Dependent Claim
37 words
The method of claim 11 , further comprising the step of applying magnetic force to isolate solid supports attached to oligonucleotides bound to said target molecules linked to magnetic beads after step c) and before step d).
Claim 15
Independent Claim
99 words
A method for identifying an aptamer which specifically binds to a target molecule other than a nucleic acid, comprising the steps of: a) providing a collection of fluorescence labeled oligonucleotides of different nucleotide sequences attached to a plurality of solid supports; b) providing a plurality of solid supports having said target molecule immobilized on their surface; c) admixing said collection of oligonucleotides with said plurality of solid supports under conditions wherein the oligonucleotide ligands become bound to the target molecule; d) isolating solid supports attached with fluorescence labeled oligonucleotides; and e) identifying oligonucleotides attached to said isolated solid supports.
Claim 16
Independent Claim
99 words
A method for identifying an aptamer which specifically binds to a target molecule other than a nucleic acid, comprising the steps of: a) providing a collection of fluorescence labeled oligonucleotides of different nucleotide sequences attached to a plurality of solid supports; b) providing a plurality of solid supports having said target molecule immobilized on their surface; c) admixing said collection of oligonucleotides with said plurality of solid supports under conditions wherein the oligonucleotide ligands become bound to the target molecule; d) isolating solid supports attached with fluorescence labeled oligonucleotides; and e) identifying oligonucleotides attached to said isolated solid supports.
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USPTO Annuity & Maintenance Windows
| Annuity Stage | Window Opens | Payment Deadline | Surcharge Period Until | Estimated Fee | Status |
|---|---|---|---|---|---|
| 3.5 Years (Year 4) | 2004-01-30 | 2004-07-31 | 2005-01-30 | $2000.00 | Window Closed / Lapsed |
| 7.5 Years (Year 8) | 2008-01-30 | 2008-07-31 | 2009-01-30 | $3760.00 | Window Closed / Lapsed |
| 11.5 Years (Year 12) | 2012-01-30 | 2012-07-31 | 2013-01-30 | $7700.00 | Window Closed / Lapsed |