Methods of localizing nucleic acids to arrays

• US 9,376,710 B2
• Filed: 01/08/2015
• Issued: 06/28/2016
• Est. Priority Date: 01/07/2004
• Status: Active Grant

• Current Assignee: Illumina Cambridge Limited (Illumina Incorporated)
• Original Assignee: Illumina Cambridge Limited (Illumina Incorporated)
• Inventors: Smith, Mark Edward Brennan, Sabot, Andrea, Rasolonjatovo, Isabelle Marie Julia, Sohna Sohna, Jean-Ernest, Horgan, Adrian Martin, Swerdlow, Harold Philip
• Primary Examiner(s): GARYU, LIANKO G
• Application Number:
  US14/592,766
• Publication Number:
  US 20150119295A1
• Time in Patent Office: 537 Days
• Field of Search: None
• US Class Current: 1/1
• 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 2219/00351 Means for dispensing and evacuation of reagents
• CPC Class Codes: B01J 2219/00527 Sheets
• CPC Class Codes: B01J 2219/00529 DNA chips
• CPC Class Codes: B01J 2219/00585 Parallel processes
• CPC Class Codes: B01J 2219/00596 Solid-phase processes
• CPC Class Codes: B01J 2219/00605 the compounds being directly bound or immobilised to solid supports
• CPC Class Codes: B01J 2219/00608 DNA chips
• CPC Class Codes: B01J 2219/0061 The surface being organic
• CPC Class Codes: B01J 2219/00612 the surface being inorganic
• CPC Class Codes: B01J 2219/00617 by chemical means
• CPC Class Codes: B01J 2219/00626 Covalent
• CPC Class Codes: B01J 2219/00637 by coating it with another layer
• CPC Class Codes: B01J 2219/00639 the compounds being trapped in or bound to a porous medium
• CPC Class Codes: B01J 2219/00641 the porous medium being continuous, e.g. porous oxide substrates
• CPC Class Codes: B01J 2219/00716 Heat activated synthesis
• CPC Class Codes: B01J 2219/0072 Organic compounds
• CPC Class Codes: B01J 2219/00722 Nucleotides
• CPC Class Codes: C08F 222/38 Amides
• CPC Class Codes: C12Q 1/6806 Preparing nucleic acids for analysis, e.g. for polymerase chain reaction [PCR] assay C12Q1/6804 takes precedence
• CPC Class Codes: C12Q 1/6834 : Enzymatic or biochemical coupling of nucleic acids to a solid phase
• CPC Class Codes: C12Q 1/6837 : using probe arrays or probe chips C12Q1/6874 takes precedence
• CPC Class Codes: C12Q 1/6876 : Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
• CPC Class Codes: C40B 40/06 : Libraries containing nucleotides or polynucleotides, or derivatives thereof
• CPC Class Codes: C40B 50/18 : using a particular method of attachment to the solid support
• CPC Class Codes: Y10T 428/265 : 1 mil or less

Abstract

Methods and compositions are disclosed relating to the localization of nucleic acids to arrays such as silane-free arrays, and of sequencing the nucleic acids localized thereby.

Claims

1. A method of localizing a target nucleic acid molecule to a solid support comprising:

providing a solid support comprising;

a surface;

a plurality of nucleic acids immobilized on the surface, wherein the nucleic acids in the plurality each comprise a primer sequence selected from SEQ ID NO;

3 or SEQ ID NO;

4;

providing the target nucleic acid molecule, wherein the target nucleic acid comprises a sequence that hybridizes to the primer sequence; and

hybridizing the target nucleic acid molecule to at least one of the plurality of nucleic acids immobilized on the surface, thereby localizing the target nucleic acid molecule to a solid support.

2. The method of claim 1, further comprising:

contacting the target nucleic acid molecule hybridized to at least one of the plurality of nucleic acids immobilized on the surface with reagents necessary to synthesize a complement of at least part of the target nucleic acid molecule, wherein the complement to at least part of the target nucleic acid molecule is covalently bound to the 3′

end of at least one of the nucleic acids immobilized on the surface.

3. The method of claim 2, further comprising separating the target nucleic acid molecule from said solid support.

4. The method of claim 3, further comprising determining the sequence of the target nucleic acid molecule.

5. The method of claim 1, wherein the surface comprises a hydrogel.

6. The method of claim 5, wherein the thickness of the hydrogel is less than 100 nm.

7. The method of claim 5, wherein the hydrogel is the product of polymerizing a first comonomer with a second comonomer, wherein the first comonomer is selected from the group consisting of acrylamide, methacrylamide, hydroxyethyl methacrylate and N-vinyl pyrrolidinone, and wherein the second comonomer is a functionalized comonomer selected from the group consisting of acrylamide, acrylate, methacrylate and methacrylamide.

8. The method of claim 1, wherein the target nucleic acid molecule comprises a sequence that is reverse-complementary to the primer sequence.

9. The method of claim 1, wherein the plurality of nucleic acids is immobilized on the surface via a linkage at their 5′

ends.

10. A composition comprising:

(a) a solid support comprising a surface; and

(b) a plurality of nucleic acids immobilized on the surface, wherein the nucleic acids in the plurality each comprise a primer sequence selected from SEQ ID NO;

3 or SEQ ID NO;

4.

11. The composition of claim 10, wherein the nucleic acids are covalently attached to a hydrogel immobilized on the surface.

12. The composition of claim 11, wherein of the hydrogel has a thickness less than 100 nm.

13. The composition of claim 11, wherein the hydrogel is the product of polymerizing a first comonomer with a second comonomer, wherein the first comonomer is selected from the group consisting of acrylamide, methacrylamide, hydroxyethyl methacrylate and N-vinyl pyrrolidinone, and wherein the second comonomer is a functionalized comonomer selected from the group consisting of acrylamide, acrylate, methacrylate and methacrylamide.

14. The composition of claim 10, wherein the nucleic acids in the plurality each comprise the primer sequence and a template, wherein the template comprises a nucleotide sequence that is longer than 300 nucleotides.

15. The composition of claim 10, wherein the nucleic acids in the plurality each comprise a polyT spacer 5′

of the primer sequence 5′

, wherein the polyT spacer comprises from 2 to 10 thymine nucleotides.

16. The composition of claim 10, wherein the solid support comprises a silica-based substrate, plastic, gold, titanium dioxide or silicon.

17. The composition of claim 10, wherein the nucleic acids in the plurality are hybridized to target nucleic acids comprising a sequence that is the reverse-complement of the primer sequence.

18. The composition of claim 10, wherein the primer sequence is at the 3′

end of the nucleic acids in the plurality.

19. The composition of claim 10, wherein the nucleic acids in the plurality further comprise a fluorescent label.

20. The composition of claim 19, wherein the nucleic acids in the plurality further comprise a 3′

blocking group.