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Details for Patent: 6,030,776

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Details for Patent: 6,030,776

Title: Parallel SELEX
Abstract:This invention discloses a method for coevolving products from two or more reactants, along with the nucleic acid that can facilitate the reaction for making the products. The invention further discloses the products and facilitating nucleic acids produced by said method.
Inventor(s): Eaton; Bruce (Boulder, CO), Gold; Larry (Boulder, CO)
Assignee: NeXstar Pharmaceuticals, Inc. (Boulder, CO)
Filing Date:Feb 24, 1997
Application Number:08/793,398
Claims:1. A method for identifying a product from a product library, wherein said product is selected for its ability to perform a preselected function on a target, said method comprising:

(a) preparing a nucleic acid-reactant test mixture comprised of nucleic acids each having a region of randomized sequence and each being coupled to a first reactant;

(b) reacting said nucleic acid-reactant test mixture with a free reactant to form a product library comprised of products formed by the reaction of said first and free reactants, wherein said reaction is facilitated by the nucleic acid coupled to said first reactant;

(c) partitioning between members of said product library based on their relative ability to perform said preselected function on a targe; and

(d) identifying said product having the ability to perform a preselected function on said target.

2. The method of claim 1 wherein said ability to perform a preselected function on a target is binding to said target.

3. The method of claim 1 wherein said ability to perform a preselected function on a target is reacting with said target and changing the property of said target.

4. The method of claim 1 wherein said nucleic acid-reactant test mixture comprises nucleic acids having a region of conserved sequences and a region of randomized sequences.

5. The method of claim 1 wherein said nucleic acid is selected from the group of single-stranded RNA, single-stranded DNA and double-stranded DNA.

6. The method of claim 1 wherein said nucleic acid-reactant test mixture comprises modified nucleotides.

7. The method of claim 6 wherein said modified nucleotides have been chemically modified at the ribose and/or phosphate and/or base positions.

8. The method of claim 6 wherein said modified nucleotides are pyrimidines modified at the 2'- or 5-positions.

9. The method of claim 6 wherein said modified nucleotides are purines modified at the 8-position.

10. The method of claim 6 wherein said modified nucleotides are modified with a chemical group which increases charge, polarizability, hydrogen bonding, electrostatic interaction or fluxionality of the nucleotide.

11. The method of claim 10 wherein said chemical group is selected from the group consisting of hydrophobic moieties, hydrophilic moieties, metal atoms in various oxidation states, rigid structures, imidazoles, primary alcohols, carboxylates, guanidium groups, amino groups, thiols and organometallic catalysts.

12. The method of claim 10 wherein said chemical group comprises an amino acid side chain or analogs thereof.

13. The method of claim 1 further comprising an organometallic catalyst incorporated into said nucleic acid test mixture.

14. The method of claim 1 which further comprises a linker group coupled between said first reactant and said nucleic acid.

15. The method of claim 14 wherein said linker group has a size in the range of 10 to 1000 .ANG..

16. The method of claim 15 wherein said linker group is selected from the group consisting of PEG, polyvinyl alcohol, polyacrylates and polypeptides.

17. The method of claim 1 wherein said first reactant is a dienophile, said free reactant is a diene, and said product is a cyclohexene derivative.

18. A method for identifying a product having the ability to perform a preselected function on a target from a product library wherein said product library is comprised of products produced from a reaction between two reactants, wherein each product is coupled to a nucleic acid that facilitates the formation of said product, said method comprising partitioning between members of said product library based on their relative ability to perform a preselected function, and identifying said product having the ability to perform said preselected function.

19. A method for identifying a product from a product library, wherein said product is selected for its ability to bind a target molecule, said method comprising:

(a) preparing a nucleic acid-reactant test mixture comprised of nucleic acids each having a region of randomized sequence and each being coupled to a first reactant;

(b) reacting said nucleic acid-reactant test mixture with a free reactant to form a product library comprised of nucleic acids coupled to a product formed by the reaction of said first and second reactants, wherein said reaction is facilitated by a nucleic acid coupled to said first reactant;

(c) contacting the product library with the target wherein products having an increased affinity to the target relative to the product library may be partitioned from the remainder of the product library;

(d) partitioning said products having increased affinity to the target from the remainder of the product library; and

(e) identifying said product having increased affinity to the target.

20. The method of claim 19 which further comprises between steps b and c, contacting the product library with a non-target and partitioning away products which bind to said non-target.

21. A method for producing a product having the ability to perform a preselected function on a target comprising:

(a) coupling each member of a nucleic acid test mixture with a first reactant to form a nucleic acid-reactant test mixture;

(b) forming a product library by contacting said nucleic acid-reactant test mixture with a mixture of free reactants, wherein said product library is formed as a result of bond formation between said first reactant and at least one of said free reactants, wherein said bond formation reaction is facilitated by a nucleic acid coupled to said first reactant;

(c) contacting the product library of step (b) with a target, wherein products having the ability to perform a preselected function on said target relative to the product library may be partitioned from the remainder of the product library; and

(d) partitioning said products having said ability to perform a preselected function of said target from the remainder of the product library, whereby said product is produced.

22. A method for identifying a facilitating nucleic acid, wherein said nucleic acid is selected for its ability to facilitate the reaction of a plurality of reactants to form a product having the ability to perform a preselected function, said method comprising:

(a) preparing a nucleic acid-reactant test mixture comprised of nucleic acids each having a region of randomized sequence and each being coupled to a first reactant;

(b) mixing said nucleic acid-reactant test mixture with a plurality of free reactants under conditions favorable for nucleic acid facilitated reaction between said reactants to form said product;

(c) isolating members of said nucleic acid-reactant test mixture which formed a product; and

(d) identifying said facilitating nucleic acid.

23. A method for identifying a product selected for its ability to perform a preselected function comprising coevolving a class of facilitating nucleic acids simultaneously with a product library, wherein said facilitating nucleic acids facilitate the formation of products from a reaction between first and free reactants, wherein each of said facilitating nucleic acids is coupled to a first reactant, and identifying said product.

24. A method for simultaneously identifying facilitating nucleic acids and producing a product library, wherein said facilitating nucleic acids facilitate the reaction between at least a first and a free reactant to form said product library, said method comprising:

(a) preparing a nucleic acid-reactant test mixture comprised of nucleic acids each having a region of randomized sequence and each being coupled to a first reactant; and

(b) reacting said nucleic acid-reactant test mixture with a free reactant to form a product library, whereby said facilitating nucleic acids can be identified.

25. A method for coproducing a facilitating nucleic acid and a product having the ability to perform a preselected function on a target comprising:

(a) coupling a first reactant to each member of a nucleic acid test mixture to form a nucleic acid-reactant test mixture;

(b) forming a product library by contacting said nucleic acid-reactant test mixture with a mixture of free reactants, wherein said product library is formed as a result of bond formation between said first reactant and at least one of said free reactants, wherein said bond formation is facilitated by a nucleic acid which is coupled to said first reactant and wherein each product is coupled to a nucleic acid;

(c) contacting said product library with a target, wherein products having the ability to perform a preselected function on said target relative to the product library may be partitioned from the remainder of the product library; and

(d) amplifying the nucleic acid coupled to the product having said ability to perform said preselected function on said target, to yield a mixture of nucleic acids enriched for facilitating nucleic acids, whereby said facilitating nucleic acid and said product having said ability to perform said preselected function on said target are produced.

26. The method of claim 25 which further comprises:

(e) coupling the facilitating nucleic acids with said first chemical reactant; and

(f) repeating steps (b) through (e) until the product having said ability to perform said preselected function on said target can be produced in sufficient quantity for structure determination.

27. A method for producing a product library comprising contacting a mixture of first reactants each coupled to a member of a nucleic acid test mixture with a mixture of, free reactants, wherein said product library is formed as a result of bond formation between said first reactant and at least one of said free reactants, wherein said bond formation reaction is facilitated by the nucleic acid coupled to said first reactant.

28. A method for producing a product comprising contacting a first reactant coupled to a facilitating nucleic acid with a free reactant, wherein said product is produced as a result of a bond formation reaction between said first reactant and at least one of said free reactants, wherein said bond formation reaction is facilitated by said facilitating nucleic acid.
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