Catalyst-free surface functionalization and polymer grafting

• US 9,994,687 B2
• Filed: 06/26/2014
• Issued: 06/12/2018
• Est. Priority Date: 07/01/2013
• Status: Active Grant

• Current Assignee: Illumina Incorporated
• Original Assignee: Illumina Incorporated
• Inventors: Berti, Lorenzo, Brown, Andrew A., George, Wayne N.
• Primary Examiner(s): Ferre, Alexandre F
• Assistant Examiner(s): Vazquez, Elaine M
• Application Number:
  US14/316,478
• Publication Number:
  US 20150005447A1
• Time in Patent Office: 1,447 Days
• Field of Search: None
• CPC Class Codes: B01J 19/0046 Sequential or parallel reactions, e.g. for the synthesis of polypeptides or polynucleotides; Apparatus and devices for combinatorial chemistry or for making molecular arrays synthesis methods per se C40B50/00
• CPC Class Codes: B01J 2219/00529 DNA chips
• CPC Class Codes: B01J 2219/00533 essentially rectangular
• CPC Class Codes: B01J 2219/00619 using hydrophilic or hydrophobic regions
• CPC Class Codes: B01J 2219/00626 Covalent
• CPC Class Codes: B01J 2219/00637 by coating it with another layer
• CPC Class Codes: B01J 2219/00722 Nucleotides
• CPC Class Codes: C08F 220/603 and containing oxygen in addition to the carbonamido oxygen and nitrogen
• CPC Class Codes: C08F 8/00 Chemical modification by after-treatment graft polymers, block polymers, crosslinking with unsaturated monomers or with polymers C08F251/00 - C08F299/00; of conjugated diene rubbers C08C
• CPC Class Codes: C08J 2335/00 Characterised by the use of homopolymers or copolymers of compounds having one or more unsaturated aliphatic radicals, each having only one carbon-to-carbon double bond, and at least one being terminated by a carboxyl radical, and containing at least one other carboxyl radical in the molecule, or of salts, anhydrides, esters, amides, imides or nitriles thereof; Derivatives of such polymers
• CPC Class Codes: C08J 7/12 Chemical modification
• CPC Class Codes: C09D 133/26 Homopolymers or copolymers of acrylamide or methacrylamide
• CPC Class Codes: C12Q 1/6837 using probe arrays or probe chips C12Q1/6874 takes precedence
• CPC Class Codes: C12Q 1/6874 involving nucleic acid arrays, e.g. sequencing by hybridisation
• CPC Class Codes: C12Q 1/6876 Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
• CPC Class Codes: C12Q 2527/125 Specific component of sample, medium or buffer
• CPC Class Codes: C12Q 2565/537 characterised by the capture oligonucleotide acting as a primer
• CPC Class Codes: C12Q 2565/629 being a microfluidic device

Abstract

Some embodiments described herein relate to a substrate with a surface comprising a silane or a silane derivative covalently attached to optionally substituted cycloalkene or optionally substituted heterocycloalkene for direct conjugation with a functionalized molecule of interest, such as a polymer, a hydrogel, an amino acid, a nucleoside, a nucleotide, a peptide, a polynucleotide, or a protein. In some embodiments, the silane or silane derivative contains optionally substituted norbornene or norbornene derivatives. Method for preparing a functionalized surface and the use in DNA sequencing and other diagnostic applications are also disclosed.

Claims

1. A substrate comprising a first surface comprising silane or a silane derivative covalently bound, optionally through a linker, to a functionalized molecule through reaction of the functionalized molecule with a first plurality of unsaturated moieties selected from the group consisting of cycloalkenes, heterocycloalkenes, and optionally substituted variants and combination thereof covalently attached to said silane or silane derivative, wherein the functionalized molecule comprises a hydrogel or a polymer that comprises a recurring unit of Formula (I) and a recurring unit of Formula (II):

2. The substrate of claim 1, wherein the first plurality of unsaturated moieties is selected from the group consisting of norbornene, heteronorbornenes, norbornene derivatives, trans-cyclooctene, trans-cyclooctene derivatives, and optionally substituted variants and combinations thereof.

3. The substrate of claim 1, wherein the first plurality of unsaturated moieties is optionally substituted norbornene.

4. The substrate of claim 1, wherein the linker is present and is covalently attached between silicon atoms of the silane or silane derivative and the first plurality of unsaturated moieties.

5. The substrate of claim 4, wherein the linker is selected from the group consisting of optionally substituted alkylenes, optionally substituted heteroalkylenes, optionally substituted cycloalkylenes, optionally substituted heterocyclylenes, optionally substituted arylenes, optionally substituted heteroarylenes, optionally substituted polyethylene glycols, cleavable linkers, and combinations thereof.

6. The substrate of claim 5, wherein the linker is selected from the group consisting of optionally substituted alkylenes and optionally substituted heteroalkylenes.

7. The substrate of claim 1, wherein R^A is selected from the group consisting of azido, optionally substituted alkenyl, optionally substituted hydrazone, optionally substituted hydrazine, and optionally substituted tetrazine.

8. The substrate of claim 1, wherein:

R⁴ is —

C(O)NH₂;

R⁵ is H or optionally substituted alkyl;

each R^4′

and R^5′

is H;

X is optionally substituted —

(CH₂)_p—

; and

p is an integer in the range of 1 to 50.

9. The substrate of claim 8, wherein p is 5 and R^A is azido.

10. The substrate of claim 1, further comprising oligonucleotides covalently attached to the functionalized molecule through a second plurality of unsaturated moieties on the oligonucleotides that are selected from the group consisting of cycloalkenes, cycloalkynes, heterocycloalkenes, heterocycloalkynes, alkynes, and optionally substituted variants and combinations thereof.

11. The substrate of claim 10, wherein the second plurality of unsaturated moieties is selected from the group consisting of optionally substituted cyclooctyne, optionally substituted cyclooctyne, optionally substituted bicyclononyne, and optionally substituted bicyclo[6.1.0] non-4-yne.

12. The substrate of claim 10, wherein the second plurality of unsaturated moieties on the oligonucleotides are alkynes or optionally substituted variants thereof.

13. The substrate of claim 1, wherein the substrate is selected from the group consisting of a glass substrate, a silica substrate, a plastic substrate, a quartz substrate, a metal substrate, a metal oxide substrate, and combinations thereof.

14. The substrate of claim 1, wherein the first surface is patterned with nanowells silanized with the silane or silane derivative, wherein the silane or silane derivative is linked to the hydrogel or polymer.

15. A method of preparing the first surface of the substrate of claim 1, said method comprising:

applying silane or the silane derivative comprising the first plurality of unsaturated moieties covalently attached thereto onto the first surface of the substrate; and

covalently attaching the functionalized molecule to said silane or silane derivative by reacting the functionalized molecule with the first plurality of unsaturated moieties to form a coating layer.

16. The method of claim 15, wherein the first plurality of unsaturated moieties of the silane or silane derivative is optionally substituted norbornene.

17. The method of claim 15, wherein the silane or silane derivative are applied onto the first surface by chemical vapor deposition.

18. The method of claim 15, further comprising

providing oligonucleotides comprising a second plurality of unsaturated moieties selected from the group consisting of cycloalkenes, cycloalkynes, heterocycloalkenes, heterocycloalkynes, alkynes, and optionally substituted variants and combinations thereof;

reacting the second plurality of unsaturated moieties of the oligonucleotides with the functionalized molecule to form covalent bonding.

19. The method of claim 18, wherein the first plurality of unsaturated moieties is selected from the group consisting of norbornene, heteronorbornenes, norbornene derivatives, trans-cyclooctene, and trans-cyclooctene derivatives, and optionally substituted variants and combinations thereof;

and wherein the second plurality of unsaturated moieties is selected from the group consisting of cyclooctyne and bicycloalkynes, and optionally substituted variants and combinations thereof.

20. The method of claim 18, wherein the second plurality of unsaturated moieties of the oligonucleotides is selected from the group consisting of optionally substituted cyclooctyne, optionally substituted bicyclononyne, and optionally substituted bicyclo[6.1.0]non-4-yne.

21. The method of claim 18, wherein the second plurality of unsaturated moieties of said oligonucleotides are alkynes or optionally substituted variants thereof.

22. The method of claim 15, wherein the silane or silane derivative comprise linkers covalently attached between silicon atoms and the first plurality of unsaturated moieties.

23. The method of claim 22, wherein the linkers are selected from the group consisting of optionally substituted alkylenes, optionally substituted heteroalkylenes, optionally substituted cycloalkylenes, optionally substituted heterocyclylenes, optionally substituted arylenes, optionally substituted heteroarylenes, optionally substituted polyethylene glycols, cleavable linkers, and combinations thereof.

24. The method of claim 22, wherein the linkers are selected from the group consisting of optionally substituted alkylenes and optionally substituted heteroalkylenes.

25. The method of claim 15, wherein:

R⁴ is —

C(O)NH₂;

R⁵ is H or optionally substituted alkyl;

each R^4′

and R^5′

is H;

X is optionally substituted —

(CH₂)_p—

; and

p is an integer in the range of 1 to 50.

26. The method of claim 25, wherein p is 5 and R^A is azido.

27. The method of claim 15, if further comprising a washing step to remove excess unbounded functionalized molecules.

28. The method of claim 15, further comprising a drying step.