Very large scale immobilized polymer synthesis

• US 6,291,183 B1
• Filed: 11/17/1999
• Issued: 09/18/2001
• Est. Priority Date: 06/07/1989
• Status: Expired due to Fees

• Current Assignee: Affymetrix, Inc. (Thermo Fisher Scientific Incorporated)
• Original Assignee: Affymetrix, Inc. (Thermo Fisher Scientific Incorporated)
• Inventors: Pirrung, Michael C., Fodor, Stephen P. A., Stryer, Lubert, Read, J. Leighton
• Primary Examiner(s): Riley, Jezia
• Application Number:
  US09/442,028
• Time in Patent Office: 671 Days
• Field of Search: 435/5, 435/6, 536/22.1, 536/23.1, 536/24.3
• 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 2219/00315 Microtiter plates
• CPC Class Codes: B01J 2219/00432 Photolithographic masks
• CPC Class Codes: B01J 2219/00434 Liquid crystal masks
• CPC Class Codes: B01J 2219/00436 Maskless processes
• CPC Class Codes: B01J 2219/00459 Beads
• CPC Class Codes: B01J 2219/00468 by manipulation of individual beads
• CPC Class Codes: B01J 2219/00475 Sheets
• CPC Class Codes: B01J 2219/005 Beads
• CPC Class Codes: B01J 2219/00527 Sheets
• CPC Class Codes: B01J 2219/00529 DNA chips
• CPC Class Codes: B01J 2219/00531 essentially square
• CPC Class Codes: B01J 2219/00585 Parallel processes
• CPC Class Codes: B01J 2219/0059 Sequential 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 2219/00619 : using hydrophilic or hydrophobic regions
• CPC Class Codes: B01J 2219/00621 : by physical means, e.g. trenches, raised areas
• 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 2219/00648 : by the use of solid beads
• CPC Class Codes: B01J 2219/00659 : Two-dimensional arrays
• CPC Class Codes: B01J 2219/00689 : using computers
• CPC Class Codes: B01J 2219/00695 : Synthesis control routines, e.g. using computer programs
• CPC Class Codes: B01J 2219/00711 : Light-directed synthesis
• CPC Class Codes: B01J 2219/00722 : Nucleotides
• CPC Class Codes: B01J 2219/00725 : Peptides
• CPC Class Codes: B01L 7/52 : with provision for submitting samples to a predetermined sequence of different temperatures, e.g. for treating nucleic acid samples amplification or hybridisation processes per se C12Q1/68; controlling sequential reactions for synthesis B01J19/0046
• CPC Class Codes: B82Y 10/00 : Nanotechnology for information processing, storage or transmission, e.g. quantum computing or single electron logic
• 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: C07C 229/14 : to carbon atoms of carbon skeletons containing rings
• CPC Class Codes: C07C 229/16 : to carbon atoms of hydrocarbon radicals substituted by amino or carboxyl groups, e.g. ethylenediamine-tetra-acetic acid, iminodiacetic acids
• CPC Class Codes: C07D 263/44 : Two oxygen atoms
• CPC Class Codes: C07D 317/62 : with hetero atoms or with carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals, directly attached to atoms of the carbocyclic ring
• CPC Class Codes: C07H 19/04 : Heterocyclic radicals containing only nitrogen atoms as ring hetero atom
• CPC Class Codes: C07H 19/10 : with the saccharide radical esterified by phosphoric or polyphosphoric acids
• 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: C07K 1/04 : on carriers C07K1/003, C07K1/006 take precedence
• CPC Class Codes: C07K 1/042 : characterised by the nature of the carrier
• CPC Class Codes: C07K 1/045 : using devices to improve synthesis, e.g. reactors, special vessels
• CPC Class Codes: C07K 1/047 : Simultaneous synthesis of different peptide species; Peptide libraries
• CPC Class Codes: C07K 1/062 : for alpha- or omega-carboxy functions
• CPC Class Codes: C07K 17/06 : attached to the carrier via a bridging agent
• CPC Class Codes: C07K 17/14 : Peptides being immobilised on, or in, an inorganic carrier
• CPC Class Codes: C07K 7/06 : having 5 to 11 amino acids
• CPC Class Codes: C08G 69/00 : Macromolecular compounds obtained by reactions forming a carboxylic amide link in the main chain of the macromolecule products obtained from isocyanates or isothiocyanates C08G18/00; polyamide-imides C08G73/14
• CPC Class Codes: C12Q 1/6809 : Methods for determination or identification of nucleic acids involving differential detection
• CPC Class Codes: C12Q 1/6816 : characterised by the detection means C12Q1/6804 takes precedence
• 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: C40B 40/06 : Libraries containing nucleotides or polynucleotides, or derivatives thereof
• CPC Class Codes: C40B 40/10 : Libraries containing peptides or polypeptides, or derivatives thereof
• CPC Class Codes: C40B 60/14 : for creating libraries
• CPC Class Codes: C40B 80/00 : Linkers or spacers specially adapted for combinatorial chemistry or libraries, e.g. traceless linkers or safety-catch linkers
• CPC Class Codes: G01N 15/1433 : using image recognition
• CPC Class Codes: G01N 21/253 : for batch operation, i.e. multisample apparatus analytical automats G01N35/00
• CPC Class Codes: G01N 21/6428 : Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes" in vivo A61B5/00; immunoassay G01N33/53
• CPC Class Codes: G01N 21/6452 : Individual samples arranged in a regular 2D-array, e.g. multiwell plates
• CPC Class Codes: G03F 7/00 : Photomechanical, e.g. photolithographic, production of textured or patterned surfaces, e.g. printing surfaces; Materials therefor, e.g. comprising photoresists; Apparatus specially adapted therefor using photoresist structures for special production processes, see the relevant places, e.g. B44C, H01L, e.g. H01L21/00, H05K
• CPC Class Codes: G03F 7/26 : Processing photosensitive materials; Apparatus therefor G03F7/12 - G03F7/24 take precedence
• CPC Class Codes: G03F 7/265 : Selective reaction with inorganic or organometallic reagents after image-wise exposure, e.g. silylation
• CPC Class Codes: G03F 7/38 : Treatment before imagewise removal, e.g. prebaking G03F7/265 takes precedence
• CPC Class Codes: G11C 13/0014 : comprising cells based on organic memory material
• CPC Class Codes: G11C 13/0019 : comprising bio-molecules
• CPC Class Codes: H01L 25/50 : Multistep manufacturing processes of assemblies consisting of devices, each device being of a type provided for in group H01L27/00 or H01L29/00 H01L21/50 takes precedence
• CPC Class Codes: H01L 2924/00 : Indexing scheme for arrangements or methods for connecting or disconnecting semiconductor or solid-state bodies as covered by H01L24/00
• CPC Class Codes: H01L 2924/0002 : Not covered by any one of groups H01L24/00, H01L24/00 and H01L2224/00
• CPC Class Codes: Y02P 20/55 : Design of synthesis routes, e.g. reducing the use of auxiliary or protecting groups
• CPC Class Codes: Y10S 435/961 : including a step of forming, releasing, or exposing the antigen or forming the hapten-immunogenic carrier complex or the antigen per se
• CPC Class Codes: Y10S 435/968 : High energy substrates, e.g. fluorescent, chemiluminescent, radioactive
• CPC Class Codes: Y10S 435/969 : Multiple layering of reactants
• CPC Class Codes: Y10S 435/973 : Simultaneous determination of more than one analyte
• CPC Class Codes: Y10S 436/807 : Apparatus included in process claim, e.g. physical support structures
• CPC Class Codes: Y10S 436/809 : Multifield plates or multicontainer arrays
• CPC Class Codes: Y10T 428/31971 : Of carbohydrate
• CPC Class Codes: Y10T 436/2575 : Volumetric liquid transfer

Abstract

A method and apparatus for preparation of a substrate containing a plurality of sequences. Photoremovable groups are attached to a surface of a substrate. Selected regions of the substrate are exposed to light so as to activate the selected areas. A monomer, also containing a photoremovable group, is provided to the substrate to bind at the selected areas. The process is repeated using a variety of monomers such as amino acids until sequences of a desired length are obtained. Detection methods and apparatus are also disclosed.

Claims

1. An array of oligonucleotides, the array comprising:

a planar solid support having at last a first surface; and

a plurality of different oligonucleotides attached to the first surface of the planar solid support at a density exceeding 1000 different oligonucleotides/cm², wherein each of the different oligonucleotides is attached to the surface of the solid support in a different known location of less than 10<sup>−

3</sup> cm², and has a different determinable sequence.

2. The array of claim 1, wherein each different oligonucleotide is from about 4 to about 20 nucleotides in length.

3. The array of claim 1, wherein each different oligonucleotide is at least 12 nucleotides in length.

4. The array of claim 1, wherein each different oligonucleotide is 2-100 nucleotides in length.

5. The array of claim 1, wherein the array comprises at least 10,000 different oligonucleotides attached to the first surface of the planar solid support.

6. The array of claim 1, wherein each of the different known locations is physically separated from each other of the known locations.

7. The array of claim 1, wherein said planar solid support is glass.

8. The array of claim 1, wherein said oligonucleotides are attached to the first surface of the planar solid support through a linker group.

9. The array of claim 1, wherein the oglionucleotides in the different known locations at least 20% pure.

10. The array of claim 1, wherein the oligonucleotides in the different known locations are at least 50% pure.

11. The array of claim 1, wherein the oligonucleotides in the different known locations are at least 80% pure.

12. The array of claim 1, wherein the oligonucleotides in the different known locations are at least 90% pure.

13. The array of claim 1, wherein said array is produced by a binary synthesis process, said process comprising the steps of:

providing a planar solid support said solid support having a plurality of compounds immobilized on a surface thereof, said compounds having protecting group coupled thereto;

deprotecting a first portion of said plurality of compounds on said surface and not a second portion of said plurality of compounds;

reacting said first portion of said plurality of compounds with a first component of said oligonucleotide;

deprotecting at least a third portion of said plurality of compounds on said surface, said third portion comprising a fraction of said first portion of said plurality of compounds;

reacting said at least third portion of said plurality of compounds with a second component of said oligonucleotide; and

optionally repeating said binary synthesis steps to produce said oligonucleotide array.

28. The array of any of claims 1, 14 or 27, wherein the different known locations are square in shape.

29. The array of any of claims 1, 14, or 27 wherein the planar solid support is glass.

34. The array of claim 1, wherein the sequences of the oligonucleotides are known.

14. An array of nucleic acids, the array comprising:

a planar solid support having at least a first surface; and

a plurality of different nucleic acids attached to the first surface of the planar solid support at a density exceeding 1000 different nucleic acids/an², wherein each of the different nucleic acids is attached to the surface of the planar solid support in a different known location of area less than 10<sup>−

3</sup> cm², has a different determinable sequence.

15. The array of claim 14, wherein each different nucleic acid is at least 20 nucleotides in length.

16. The array of claim 14, wherein the array comprises at least 1,000 different nucleic acids attached to the first surface of the planar solid support.

17. The array of claim 14, wherein the array comprises at least 10,000 different nucleic acids attached to the first surface of the planar solid support.

18. The array of claim 14, wherein each of the different known locations is physically separated from each of the other known locations.

19. The array of claim 14, wherein said planar solid support is glass.

20. The of claim 14, wherein amid nucleic acids are attached to the first surface of the planar solid support through a linker group.

21. The array of claim 14, wherein the nucleic acids in the different known locations comprise nucleic acids that are at least 20% pure.

22. The array of claim 14, wherein the nucleic acid in the different known locations comprise nucleic acids that are at least 50% pure.

23. The array of claim 14, wherein the nucleic acids in the different known locations we at least 80% pure.

24. The array of claim 14, the nucleic acids in the different known locations are at least 90% pure.

25. The array of claim 14, wherein said array is produced by a binary synthesis process, said process comprising the steps of:

providing a planar, solid support, said solid support having a plurality of compounds immobilized on a surface thereof, said compounds having protecting groups coupled thereto;

deprotecting a first portion of said plurality of compounds on said surface and not a second portion of said plurality of compounds;

reacting said first portion of said plurality of compounds with a first reactant;

deprotecting at lest a third portion of said plurality of compounds on said surface, said third portion comprising a fraction of said first portion of said plurality of compounds;

reacting said at least third portion of said plurality of compounds with a second reactant; and

optionally repeating said binary synthesis steps to produce said nucleic acid array.

26. The array of claim 14, wherein the nucleic acids are covalently attached to the support.

30. The array of claim 14, wherein the substrate is silicon dioxide.

31. The array of claim 14, wherein the planar solid support is (poly)tetrafluroethylene, (poly)vinylidenedifluoride, polystryene or polycarbonate.

32. The array of claim 14, wherein the planar solid support is optically transparent.

33. The may of claim 14, wherein the planar solid support is functionalized with groups that attach to the plurality of different nucleic acids.

35. The array of claim 14, wherein the sequences of the nucleic acids are known.

27. An array of nucleic acids, the array comprising:

a planar solid support having at least a first surface; and

a plurality of different nucleic acids attached to the first surface of the planar solid support at a density exceeding 1000 different nucleic acids/cm², wherein each of the different nucleic acids is attached lo the surface of the solid support in a different known location of less than 10<sup>−

3</sup> cm², has a different determinable sequence, wherein the surface and the support are made from different materials.

36. The array of claim 27, wherein the sequences of nucleic acids are known.