Details for Patent: 6,465,189

Title:	Systematic evolution of ligands by exponential enrichment: blended selex
Abstract:	A method is described for generating blended nucleic acid ligands containing non-nucleic acid functional units. Specifically, a SELEX identified RNA ligand to the integrin gpIIbIIIa is conjugated to the peptide Gly-Arg-Gly-Asp-Thr-Pro (SEQ ID NO:1). This blended RNA ligand inhibits the biological activity of gpIIbIIIa with high specificity. Also described is a single-stranded DNA ligand to elastase coupled to N-methoxysuccinyl-Ala-Ala-Pro-Val-chloromethyl ketone (SEQ ID NO:2). This elastase blended nucleic acid ligand inhibits the biological activity of elastase.
Inventor(s):	Biesecker; Greg (Boulder, CO), Jayasena; Sumedha (Boulder, CO), Gold; Larry (Boulder, CO), Smith; Drew (Boulder, CO), Kirschenheuter; Gary (Arvada, CO)
Assignee:	Gilead Sciences, Inc. (Foster City, CA)
Filing Date:	Jun 29, 2000
Application Number:	09/606,477
Claims:	1. A blended nucleic acid ligand of a target compound, wherein the nucleic acid ligand is identified from a blended candidate mixture comprised of blended nucleic acids each having at least one nucleic acid region and at least one functional unit, according to the method comprising: a) contacting the blended candidate mixture with the target, wherein blended nucleic acids having an increased affinity to the target relative to the blended candidate mixture may be partitioned from the remainder of the blended candidate mixture; b) partitioning the increased affinity blended nucleic acids from the remainder of the blended candidate mixture; and c) amplifying the increased affinity blended nucleic acids to yield a ligand-enriched mixture of blended nucleic acids, whereby a blended nucleic acid ligand of the target compound may be identified. 2. A diagnostic reagent comprising the blended nucleic acid ligand of claim 1. 3. The diagnostic reagent of claim 2 wherein said at least one functional unit is a reporter molecule. 4. The diagnostic reagent of claim 3, wherein the reporter molecule is a post-method modification. 5. The diagnostic reagent of claim 3, wherein the reporter molecule is biotin. 6. The diagnostic reagent of claim 3, wherein the reporter molecule is a fluorescent tagged oligonucleotide. 7. A method for identifying a diagnostic reagent of a target from a blended candidate mixture comprised of blended nucleic acids each having at least one nucleic acid region and at least one functional unit, said method comprising: a) contacting the blended candidate mixture with the target, wherein blended nucleic acids having an increased affinity to the target relative to the blended candidate mixture may be partitioned from the remainder of the blended candidate mixture; b) partitioning the increased affinity blended nucleic acids from the remainder of the blended candidate mixture; c) amplifying the increased affinity blended nucleic acids to yield a ligand-enriched mixture of blended nucleic acids, whereby a diagnostic reagent of a target may be identified. 8. The method of claim 7 further comprising adding a reporter molecule to the diagnostic reagent. 9. The method of claim 7 wherein said at least one nucleic acid region is composed of a fixed region and a randomized region. 10. The method of claim 9 wherein said at least one functional unit is attached to an oligonucleotide hybridized to said fixed region. 11. A diagnostic reagent identified according to the method of claim 7. 12. A method for preparing a diagnostic reagent, wherein said diagnostic reagent comprises a blended nucleic acid ligand of a target compound and a reporter molecule, comprising: a) identifying a nucleic acid ligand of a target compound from a candidate mixture comprised of nucleic acids each having a region of randomized sequence by a method comprising: i) contacting the candidate mixture with the target, wherein nucleic acids having an increased affinity to the target relative to the candidate mixture; ii) partitioning the increased affinity nucleic acids from the remainder of the candidate mixture, and iii) amplifying the increased affinity nucleic acids to yield a ligand-enriched mixture of nucleic acids, whereby a nucleic acid ligand of the target compound may be identified; b) attaching at least one functional unit to said nucleic acid ligand to yield a blended nucleic acid ligand of the target compound; and c) attaching at least one reporter molecule to said blended nucleic acid ligand, whereby a diagnostic reagent is prepared.