MICROARRAY FABRICATION SYSTEM AND METHOD

• US 20130116153A1
• Filed: 10/26/2012
• Published: 05/09/2013
• Est. Priority Date: 10/28/2011
• Status: Active Grant

• Current Assignee: Illumina Incorporated
• Original Assignee: Illumina Incorporated
• Inventors: Lin, Shengrong, Vijayan, Kandaswamy, Wu, Yir-Shyuan, Venkatesan, Bala Murali, Park, Sang Ryul, Rogert Bacigalupo, Maria Candelaria, Beierle, John M., Gunderson, Kevin L., Berti, Lorenzo, Tsay, James, Bowen, M. Shane
• Granted Patent: US 8,778,849 B2
• Time in Patent Office: Days
• US Class Current: 506/16
• 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 2219/00608 DNA chips
• CPC Class Codes: B01J 2219/00621 by physical means, e.g. trenches, raised areas
• CPC Class Codes: B01J 2219/00637 by coating it with another layer
• CPC Class Codes: B01J 2219/00641 the porous medium being continuous, e.g. porous oxide substrates
• CPC Class Codes: B01J 2219/00644 the porous medium being present in discrete locations, e.g. gel pads
• CPC Class Codes: B01J 2219/00648 by the use of solid beads
• CPC Class Codes: B01J 2219/00653 the compounds being bound to electrodes embedded in or on the solid supports
• CPC Class Codes: B01J 2219/00659 Two-dimensional arrays
• CPC Class Codes: B01J 2219/00662 Two-dimensional arrays within two-dimensional arrays
• CPC Class Codes: B01J 2219/00722 Nucleotides
• CPC Class Codes: B01J 2219/00788 Three-dimensional assemblies, i.e. the reactor comprising a form other than a stack of plates
• CPC Class Codes: C12Q 1/6844 Nucleic acid amplification reactions
• CPC Class Codes: C12Q 1/686 Polymerase chain reaction [PCR]
• CPC Class Codes: C12Q 2527/125 Specific component of sample, medium or buffer
• CPC Class Codes: C12Q 2535/122 Massive parallel sequencing
• CPC Class Codes: C12Q 2565/543 characterised by the use of two or more capture oligonucleotide primers in concert, e.g. bridge amplification
• 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

Abstract

A microarray is designed capture one or more molecules of interest at each of a plurality of sites on a substrate. The sites comprise base pads, such as polymer base pads, that promote the attachment of the molecules at the sites. The microarray may be made by one or more patterning techniques to create a layout of base pads in a desired pattern. Further, the microarrays may include features to encourage clonality at the sites.

Claims

1. A method for preparing a biological microarray, comprising:

providing an array of base pads at predetermined sites on a substrate, wherein individual base pads are configured to capture a nucleic acid molecule; and

contacting the array of base pads with a mixture of different nucleic acid molecules under conditions wherein a nucleic acid molecule is captured at each base pad,

wherein a porous attachment layer is disposed over the base pads and the porous attachment layer is configured to attach amplified copies of the nucleic acid molecules comprising nucleotides or nucleotide-like components.

2. The method of claim 1, wherein individual base pads are configured to capture no more than one nucleic acid molecule.

3. The method of claim 2, wherein a single nucleic acid molecule is captured by each and every base pad in the array.

4. The method of claim 2, wherein a single nucleic acid molecule is captured by fewer than all of the base pads in the array.

5. The method of claim 1, wherein the nucleic acid molecules comprise nucleotides or nucleotide-like components.

6. The method of claim 1, further comprising amplifying the single nucleic acid molecule at each base pad to obtain at each base pad a region comprising copies of the nucleic acid molecule, wherein the copies of the nucleic acid molecule are attached to the porous attachment layer.

7. The method of claim 1, wherein the substrate comprises a glass.

8. The method of claim 1, wherein the base pads comprise gold.

9. The method of claim 1, wherein the base pads are formed by an imprint lithographic process.

10. The method of claim 9, wherein the imprint lithographic process comprises disposing a transfer layer on the substrate, disposing a light-curing resist layer over the transfer layer, patterning the resist layer via a mold, curing the patterned resist layer, etching the resist and transfer layers to expose regions of the substrate, depositing a pad-forming substance over the exposed regions of the substrate, and removing the resist and transfer layers.

11. The method of claim 10, wherein curing the patterned resist layer comprises exposing the mold to radiation that traverses the mold to cure the patterned resist layer without removing the mold from the patterned resist layer.

12. The method of claim 1, wherein the porous attachment layer is patterned.

13. The method of claim 12, wherein the porous attachment layer comprises a plurality of regions at least partially surrounding each of the base pads.

14. The method of claim 1, wherein the porous attachment layer extends substantially continuously over the plurality of base pads.

15. The method of claim 1, wherein the porous attachment layer comprises acrylamide.

16. A method for preparing a biological microarray, comprising:

forming an array of base pads at predetermined sites on a substrate;

disposing a molecule binding substance over each of the base pads, thereby configuring each of the base pads to capture a nucleic acid molecule;

disposing a porous attachment layer over the base pads;

seeding each of the base pads with a single nucleic acid molecule by linking the single nucleic acid molecule to the molecule binding substance; and

amplifying the nucleic acid molecule at each base pad to obtain at each base pad a region comprising copies of the nucleic acid molecule, wherein the copies of the nucleic acid molecule are attached to the porous attachment layer.

17. A biological microarray system, comprising:

an array of base pads at predetermined sites on a substrate;

a molecule binding substance disposed over each of the base pads and linked to no more than a nucleic acid molecule;

a porous attachment layer disposed over the base pads; and

several copies of each of the nucleic acid molecules linked to the porous attachment layer disposed over each of the respective base pads.

18. A method for preparing a biological microarray, comprising:

forming a polymer layer on a substrate;

disposing a photoresist layer over the polymer layer;

forming interstitial spaces in the photoresist layer and the polymer layer;

removing the photoresist layer to expose polymer base pads, wherein the polymer base pads are coupled to a molecule binding substance.

19. The method of claim 18, wherein the polymer layer comprises a poly(N-(5-azidoacetamidylpentyl)acrylamide-co-acrylamide) polymer.

20. A method for preparing a biological microarray, comprising:

activating regions on a substrate to form a pattern of activated regions;

contacting the substrate with a self-assembling monomer solution;

polymerizing the monomers to form polymer base pads only on the activated regions wherein the polymer base pads are coupled to a molecule binding substance.

21. A method for preparing a biological microarray, comprising:

forming an electrically conductive layer on a surface of a substrate;

forming a plurality of spaced apart electrically nonconductive regions on the electrically conductive layer;

forming a polymer layer over the electrically conductive layer and the plurality of spaced apart electrically nonconductive regions, wherein the polymer is coupled to a plurality of primers; and

applying a current through the electrically conductive layer to deactivate only a portion of the primers coupled to the polymer.

22. A method for preparing a biological microarray, comprising:

forming a polymer layer on a surface of a substrate;

forming a plurality of spaced apart photoresist regions on the polymer;

contacting exposed portions of the polymer layer with a plurality of primers; and

removing the photoresist regions and covered portions of the polymer layer such that a plurality of spaced apart polymer pads coupled the plurality of primers remain on the surface of the substrate.

23. A method for preparing a biological microarray, comprising:

forming wells on a substrate, wherein the wells are separated by photoresist interstitial regions;

applying nanoparticles on the substrate such that the nanoparticles cover the wells and the photoresist interstitial regions;

removing the photoresist such that a plurality of spaced apart nanoparticles remain on the surface of the substrate; and

coupling a molecule binding substance to the nanoparticles.

24. A method for amplifying nucleic acids, comprising

(a) providing an amplification reagent comprising

(i) an array of amplification sites, and

(ii) a solution comprising a plurality of different target nucleic acids,

wherein the different target nucleic acids have fluidic access to the plurality of amplification sites and wherein the solution comprises at least 3% PEG, and

(b) reacting the amplification reagent to produce a plurality of amplification sites that each comprise a clonal population of amplicons from an individual target nucleic acid from the solution, wherein the reacting comprises (i) producing a first amplicon from an individual target nucleic acid that transports to each of the amplification sites, and (ii) producing subsequent amplicons from the individual target nucleic acid that transports to each of the amplification sites or from the first amplicon.

25. A method for amplifying nucleic acids, comprising

(a) providing an amplification reagent in a flow cell comprising

(i) an array of amplification sites, and

(ii) a solution comprising a plurality of different target nucleic acids,

wherein the different target nucleic acids have fluidic access to the plurality of amplification sites,

(b) appling an electric field across the flow cell to crowd the target nucleic acids towards the array of amplification sites;

and

(c) reacting the amplification reagent to produce a plurality of amplification sites that each comprise a clonal population of amplicons from an individual target nucleic acid from the solution, wherein the reacting comprises (i) producing a first amplicon from an individual target nucleic acid that transports to each of the amplification sites, and (ii) producing subsequent amplicons from the individual target nucleic acid that transports to each of the amplification sites or from the first amplicon.