Use of ionic liquids for fabrication of polynucleotide arrays

• US 20050112679A1
• Filed: 12/22/2004
• Published: 05/26/2005
• Est. Priority Date: 10/31/2001
• Status: Active Grant

• Current Assignee: Douglas J. Delinger, Geraldine F. Dellinger, Joel Myerson, Michel G.M. Perbost
• Original Assignee: Douglas J. Delinger, Geraldine F. Dellinger, Joel Myerson, Michel G.M. Perbost
• Inventors: Delinger, Douglas J., Dellinger, Geraldine F., Myerson, Joel, Perbost, Michel G. M.
• Granted Patent: US 7,435,810 B2
• Time in Patent Office: Days
• US Class Current: 506/16
• CPC Class Codes: B01J 2219/00378 Piezoelectric or ink jet dispensers
• CPC Class Codes: B01J 2219/00608 DNA chips
• CPC Class Codes: B01J 2219/00612 the surface being inorganic
• CPC Class Codes: B01J 2219/00626 Covalent
• CPC Class Codes: B01J 2219/00675 In-situ synthesis on the substrate
• CPC Class Codes: B01J 2219/00677 Ex-situ synthesis followed by deposition on the substrate
• CPC Class Codes: B82Y 30/00 Nanotechnology for materials or surface science, e.g. nanocomposites
• CPC Class Codes: C07B 2200/11 Compounds covalently bound to a solid support
• CPC Class Codes: C07H 21/00 Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids
• CPC Class Codes: C40B 50/14 Solid phase synthesis, i.e. wherein one or more library building blocks are bound to a solid support during library creation; Particular methods of cleavage from the solid support
• CPC Class Codes: C40B 60/14 for creating libraries
• CPC Class Codes: Y02P 20/54 using solvents, e.g. supercritical solvents or ionic liquids

Abstract

A method of fabricating polynucleotide arrays includes dissolving a nucleotide monomer, oligonucleotide, or polynucleotide in a solvent containing ionic liquid and depositing the resulting solution on an array substrate. The method has particular application to fabrication of an addressable array of polynucleotides on a substrate that carries substrate bound moieties each with a hydroxyl group. The process may be repeated at specific locations on the array to elongate the polynucleotide deposited on the array.

Claims

1. A method of immobilizing a nucleotide monomer, oligonucleotide, or polynucleotide upon an insoluble substrate, the method comprising

a) dissolving a nucleotide monomer, oligonucleotide, or polynucleotide in a solvent comprising an ionic liquid such that the resulting solution comprises at least 50% by weight ionic liquid, and

b) contacting the insoluble substrate with the solution resulting from a) under conditions and for a time sufficient to immobilize the nucleotide monomer, oligonucleotide, or polynucleotide upon the insoluble substrate.

2. The method of claim 1 wherein the solution includes at least about 75% by weight of an ionic liquid.

3. The method of claim 1 wherein the solution includes at least about 90% by weight of an ionic liquid.

4. The method of claim 1 wherein the solution includes at least about 98% by weight of an ionic liquid.

5. The method of claim 1 wherein the ionic liquid is an organic salt comprising a substituted heterocyclic organic cation.

6. The method of claim 5 wherein the ionic liquid is an organic salt comprising a cation selected from an N-substituted pyridine and a 1,3-disubstituted imidazole.

7. The method of claim 5 wherein the organic salt comprises an anion selected from chloride (Cl⁻

), bromide (Br⁻

), tetrafluoroborate ([BF₄]⁻

), hexafluorophosphate ([PF₆]⁻

), [SbF₆]⁻

, [CuCl₂]⁻

, [AlCl₄]⁻

, [Al₂Cl₇]⁻

, [Al₃Cl₁₀]⁻

, methylsulfate (CH₃SO₄⁻

), trifluoroacetate (CF₃CO₂⁻

), heptafluorobutanoate (CF₃(CF₂)₂CO₂⁻

), triflate (CF₃SO₂⁻

), nonaflate (C₂F₅SO₂⁻

), bis(trifluoromethylsulfonyl)imide ((CF₃SO₂)₂N⁻

), bis(perfluoroethylsulfonyl)imide ((C₂F₅SO₂)₂N⁻

), and tris(trifluoromethylsulfonyl)methide ((CF₃SO₂)₃C⁻

).

8. The method of claim 5 wherein the organic salt is characterized as being a liquid when being >

98% pure and at standard temperature and pressure.

9. The method of claim 5 wherein the organic salt is characterized as being a liquid below about 80°

C. at about standard atmospheric pressure.

10. The method of claim 5 wherein the cation is an N-substituted pyridine having the formula

11. The method of claim 5 wherein the cation has the formula

12. The method of claim 1 further comprising conducting steps a) and b) at each of a plurality of sites on the insoluble substrate to form an addressable array of polynucleotides on the insoluble substrate.

13. The method of claim 1 wherein the contacting is performed by depositing droplets of the solution resulting from a) on the substrate using an ink jet device.

14. A method of forming a bond between a reactive group immobilized on an insoluble substrate and a molecule selected from the group consisting of a nucleotide monomer, an olignucleotide, and a polynucleotide;

the method comprising contacting the reactive group with a solution comprising the molecule and at least 50% by weight of an ionic liquid ionic liquid.

15. The method of claim 14 wherein the solution includes at least about 75% by weight of an ionic liquid.

16. The method of claim 14 wherein the solution includes at least about 90% by weight of an ionic liquid.

17. The method of claim 14 wherein the solution includes at least about 98% by weight of an ionic liquid.

18. The method of claim 14 wherein the bond is an internucleotide bond.

19. The method of claim 18, further comprising contacting the internucleotide bond with an oxidizing reagent to oxidize the internucleotide bond.

20. The method of claim 19 wherein the insoluble substrate is a planar substrate having a surface, an array of polynucleotides being arranged upon the surface.

21. The method of claim 14 wherein the ionic liquid is an organic salt comprising a substituted heterocyclic organic cation.

22. The method of claim 14 wherein the ionic liquid is an organic salt comprising a cation selected from an N-substituted pyridine and a 1,3-disubstituted imidazole.

23. The method of claim 21, the organic salt comprising an anion selected from chloride (Cl⁻

), bromide (Br⁻

), tetrafluoroborate ([BF₄]⁻

), hexafluorophosphate ([PF₆]⁻

), [SbF₆]⁻

, [CuCl₂]⁻

, [AlCl₄]⁻

, [Al₂Cl₇]⁻

, [Al₃Cl₁₀]⁻

, methylsulfate (CH₃SO₄⁻

), trifluoroacetate (CF₃CO₂⁻

), heptafluorobutanoate (CF₃(CF₂)₂CO₂⁻

), triflate (CF₃SO₂⁻

), nonaflate (C₂F₅SO₂⁻

), bis(trifluoromethylsulfonyl)imide ((CF₃SO₂)₂N⁻

), bis(perfluoroethylsulfonyl)imide ((C₂F₅SO₂)₂N⁻

), and tris(trifluoromethylsulfonyl)methide ((CF₃SO₂)₃C⁻

).

24. The method of claim 14, the organic salt having the characteristic of being a liquid when being >

98% pure and at standard temperature and pressure.

25. The method of claim 21 wherein the cation is an N-substituted pyridine having the formula

26. The method of claim 21 wherein the cation has the formula

27. The method of claim 14 wherein the contacting is performed by depositing droplets of the solution on the substrate using an ink jet device.

28. The method of claim 14 wherein the organic salt is characterized as being a liquid below about 80°

C. at about standard atmospheric pressure