Using populations of beads for the fabrication of arrays on surfaces

• US 8,198,028 B2
• Filed: 07/02/2009
• Issued: 06/12/2012
• Est. Priority Date: 07/02/2008
• Status: Active Grant

• Current Assignee: Illumina Cambridge Limited (Illumina Incorporated)
• Original Assignee: Illumina Cambridge Limited (Illumina Incorporated)
• Inventors: Rigatti, Roberto, Smith, Geoffrey Paul, Boutell, Jonathan Mark
• Primary Examiner(s): Horlick, Kenneth R.
• Application Number:
  US12/497,397
• Publication Number:
  US 20100022412A1
• Time in Patent Office: 1,076 Days
• Field of Search: 435/6.12, 435/91, 435/1, 435/91.2
• US Class Current: 435/6.12
• 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 2219/00608 DNA chips
• CPC Class Codes: C12N 15/1068 Template (nucleic acid) mediated chemical library synthesis, e.g. chemical and enzymatical DNA-templated organic molecule synthesis, libraries prepared by non ribosomal polypeptide synthesis [NRPS], DNA/RNA-polymerase mediated polypeptide synthesis
• 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 2563/149 Particles, e.g. beads
• CPC Class Codes: C12Q 2563/161 Vesicles, e.g. liposome
• CPC Class Codes: C12Q 2565/537 characterised by the capture oligonucleotide acting as a primer
• CPC Class Codes: C12Q 2565/543 characterised by the use of two or more capture oligonucleotide primers in concert, e.g. bridge amplification

Abstract

The present invention provides methods for creating an array of features on a surface based on content transferred from a plurality of beads to the surface. Nucleic acid content can be transferred using a method including the steps of (a) providing a surface having one or more primer oligonucleotides attached to the surface; (b) providing a pool of beads, wherein beads in the pool have a plurality of templates attached thereto, the plurality comprising multiple copies of a single nucleic acid template sequence; (c) arraying the beads onto the surface by hybridizing the templates to the primer oligonucleotides; and (d) extending the primers to produce copies of the templates attached to the surface.

Claims

1. A method for fabricating an array of nucleic acids on a surface comprising:

(a) providing a surface comprising one or more primer oligonucleotides attached to the surface;

(b) providing a pool of beads, wherein beads in the pool have a plurality of templates attached thereto, the plurality comprising multiple copies of a single nucleic acid template sequence;

(c) arraying the beads onto the surface by hybridizing the templates to the primer oligonucleotides; and

(d) extending the primers to produce copies of the templates attached to the surface.

2. The method according to claim 1, wherein step (b) comprises forming the pool of beads by amplifying the nucleic acid template sequence on the beads, thereby producing the plurality of templates.

3. The method according to claim 2, wherein the amplifying is performed in an emulsion.

4. The method according to claim 2, wherein the amplifying is performed by bridge amplification using two or more primer oligonucleotides immobilized on the beads.

5. The method according to claim 4, wherein one or more of the primer oligonucleotides is cleaved after the amplification to leave single stranded templates on the beads.

6. The method of claim 1, wherein a subset of at most 10% of the beads in the pool comprise the copies of the single nucleic acid template sequence.

7. The method of claim 1, wherein a subset of at most 1% of the beads in the pool comprise the copies of the single nucleic acid template sequence.

8. The method according to claim 2, wherein a subset of at least 90% of the beads on the surface carry one or more copies of the single nucleic acid template sequence.

9. The method according to claim 2, wherein a subset of at least 99% of the beads on the surface carry one or more copies of the single nucleic acid template sequence.

10. The method according to claim 1, wherein the copies of the templates attached to the surface that are produced in step (d) comprise a plurality of nucleic acid features on the surface, wherein each of the features covers less area of the surface than the area covered by each of the beads.

11. The method according to claim 1, further comprising removing the beads from the surface after the extending of the primers.

12. The method according to claim 2, wherein the templates amplified on the beads originate from a pool of chemically synthesized oligonucleotides.

13. The method according to claim 1 further comprising an additional step of sequencing the copies of the templates attached to the surface that are produced in step (d).

14. The method of claim 1, wherein the primer oligonucleotides are attached to spatially random locations on the surface.

15. The method of claim 1, wherein the primer oligonucleotides are attached to spatially ordered locations on the surface.

16. The method of claim 1, wherein the primer oligonucleotides are attached to the surface in a number per unit area that exceeds the number of the beads arrayed within the unit area of the surface.

17. The method according to claim 1 wherein in step (d) the plurality of templates attached to the beads are each copied two or more times.

18. The method of claim 1, wherein the copies of the templates attached to the surface that are produced in step (d) comprise a plurality of nucleic acid features on the surface, and wherein the center-to-center spacing of the features is less than 5 μ

m.

19. The method of claim 1, wherein the surface is flat or planar.

20. The method of claim 1, further comprising an additional step of amplifying the copies of the templates attached to the surface.

21. The method of claim 20, wherein double stranded copies of the templates are produced by the amplifying.

22. The method of claim 1, wherein the plurality of templates attached to the beads are produced from a genomic DNA sample, RNA sample or cDNA sample.

23. The method of claim 1, wherein the plurality of templates attached to the beads are chemically synthesized oligonucleotides.

24. The method of claim 2, wherein two primers are used for the amplifying.

25. The method of claim 24, wherein the primers are complementary to universal amplification sequences at the ends of nucleic acid molecules that comprise the template sequence.

26. The method of claim 11, further comprising an additional step of hybridizing target nucleic acids to the copies of the templates attached to the surface after removing the beads from the surface, thereby detecting the target nucleic acids.

27. The method of claim 11, further comprising an additional step of amplifying the copies of the templates attached to the surface after removing the beads from the surface.

28. The method of claim 11, further comprising an additional step of sequencing the copies of the templates attached to the surface after removing the beads from the surface.

29. The method of claim 11, further comprising an additional step of obtaining an image of the surface after removing the beads from the surface.