Polypeptide arrays

• US 6,919,211 B1
• Filed: 09/01/2000
• Issued: 07/19/2005
• 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): Ponnaluri, Padmashri
• Application Number:
  US09/653,761
• Time in Patent Office: 1,782 Days
• Field of Search: 436/518, 436/528, 436/527, 436/807, 435/4, 435/DIG.22, 435/DIG.34, 435/DIG.35, 435/DIG.49, 435/6, 530/333, 530/334
• US Class Current: 506/18
• 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 2219/00621 : 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 2219/00641 : 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 2219/00659 : 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: 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 1/042 : 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 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: 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 1/6827 : for detection of mutation or polymorphism
• 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: G01N 15/1433 : using image recognition
• 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 7/265 : 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: Y10S 436/807 : Apparatus included in process claim, e.g. physical support structures

Abstract

The present invention provides methods and apparatus for sequencing, fingerprinting and mapping biological macromolecules, typically biological polymers. The methods make use of a plurality of sequence specific recognition reagents which can also be used for classification of biological samples, and to characterize their sources.

Claims

1. A substrate with a surface comprising a plurality of polypeptides with different, known sequences bound to the surface in positionally defined locations, at a density exceeding 400 different polypeptides occupying a total area of less than 1 cm² on said substrate, each of said polypeptides having a different polypeptide sequence.

2. The substrate as recited in claim 1, wherein said substrate comprises 10³ or more different polypeptides with known sequences bound to positionally defined locations of said substrate.

3. The substrate as recited in claim 1, wherein said substrate comprises 10⁴ or more different polypeptides with known sequences bound to positionally defined locations of said substrate.

4. The substrate as recited in claim 1, wherein said substrate comprises 10⁵ or more different polypeptides with known sequences bound to positionally defined locations.

5. The substrate as recited in claim 1, wherein said substrate comprises 10⁶ or more different polypeptides with known sequences bound to positionally defined locations.

6. The substrate as recited in claim 1, wherein said polypeptides are at least 50% pure within said positionally defined locations.

7. The substrate as recited in claim 1, wherein the polypeptides are covalently attached to the surface.

8. The substrate as recited in claim 1, wherein the polypeptides are attached to the surface by a linker.

9. A substrate with a surface comprising a plurality of polypeptides with different, known sequences bound to the surface in positionally defined locations, at a density exceeding 1000 different polypeptides occupying a total area of less than 1 cm² on said substrate, each of said polypeptides having a different polypeptide sequence.

10. The substrate as recited in claim 9, wherein said substrate comprises 10³ or more different polypeptides with known sequences bound to positionally defined locations of said substrate.

11. The substrate as recited in claim 9, wherein said substrate comprises 10⁴ or more different polypeptides with known sequences bound to positionally defined locations of said substrate.

12. The substrate as recited in claim 9, wherein said substrate comprises 10⁶ or more different polypeptides with known sequences bound to positionally defined locations.

13. The substrate as recited in claim 9, wherein said substrate comprises 10⁶ or more different polypeptides with known sequences bound to positionally defined locations.

14. An array of more than 1,000 different polypeptides with known sequences bound to a surface of a substrate, said polypeptides each in positionally defined locations and differing from other polypeptides in monomer sequence, each of said positionally defined locations being an area of less than about 0.01 cm² and each positionally defined location comprising polypeptides of known sequence, said different polypeptides occupying a total area of less than 1 cm².

15. The array as recited in claim 14, wherein said positionally defined locations each have an area of about 1×

10<sup>−

2</sup> cm² to about 1×

10<sup>−

5</sup> cm².

16. The array as recited in claim 15, wherein said positionally defined locations each have an area of about 1×

10<sup>−

2</sup> cm² to about 1×

10<sup>−

4</sup> cm².

17. The array as recited in claim 16, wherein said positionally defined locations each have an area of about 1×

10<sup>−

2</sup> cm² to about 1×

10<sup>−

3</sup> cm².

18. The array as recited in claim 14, made by the process of:

a) providing a polypeptide array comprising at least two different polypeptides immobilized on a surface, and wherein said polypeptides are synthesized on said surface;

b) contacting said surface with a first protected amino acid wherein said first protected amino acid is selectively coupled to a functional group in a first selectively activated region of said surface;

c) contacting said surface with a second protected amino acid without physical segregation of said surface such that said second protected amino acid is selectively coupled to a functional group in a second selectively activated region of said surface; and

d) repeating the above steps until at least 1,000 different polypeptides are formed at positionally defined locations within the selectively activated regions on said substrate surface.

19. The array as recited in claim 18, wherein said first selectively activated region of said substrate is exposed to light to remove a photoremovable group from said first protected amino acid.

20. The array as recited in claim 14, comprising more than 10,000 polypeptides of known sequences.

21. An array of polypeptides, said array of polypeptides comprising:

a substrate having a surface; and

a plurality of different polypeptides bound to said surface at a density exceeding 400 different polypeptides/cm², wherein each of said plurality of different polypeptides is attached to said surface in a different positionally defined location of area greater than 100 square, microns and has a different determinable sequence.

22. The array of claim 21, wherein said substrate is a solid support.

23. The array of claim 22, wherein said solid support is a member selected from the group consisting of particles, strands, precipitates, gels, sheets, tubing, spheres, containers, capillaries, pads, slices, films, plates and slides.

24. The array of claim 22, wherein said solid support is made of a member selected from the group consisting of polymers, plastics, resins, polysaccharides, silica or silica-based materials, carbon, metals, inorganic glasses, and membranes.

25. The array of claim 22, wherein said solid support is glass.

26. The array of claim 22, wherein said solid support is a gel.

27. The array of claim 22, wherein said polypeptides are attached to said solid support through a linker group.

28. The array of claim 22, wherein said array comprises at least 1,000 different polypeptides attached to said solid support.

29. The array of claim 22, wherein said array comprises at least 10,000 different polypeptides attached to said solid support.

30. The array of claim 22, wherein said plurality of different polypeptides attached to said surface are at a density exceeding 1000 different polypeptides/cm².

31. The array of claim 21, wherein each of said different positionally defined locations is physically separated from each of the other positionally defined locations.

32. The array of claim 21, wherein said polypeptides in said different positionally defined locations are at least 20% pure.

33. The array of claim 21, wherein said polypeptides in said different positionally defined locations are at least 50% pure.

34. The array of claim 21, wherein said polypeptides in said different positionally defined locations are at least 80% pure.

35. The array of claim 21, wherein said polypeptides in said different positionally defined locations are at least 90% pure.

36. The array of claim 22, wherein said array is produced by a process comprising:

providing a planar, non-porous 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 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 third portion of said plurality of compounds with a second reactant; and

optionally repeating said synthesis steps to produce said polypeptide array.

37. The array of claim 21, wherein said polypeptides in said different positionally defined locations are at least 10% pure.

38. The array of claim 21, wherein said substrate is rigid.

39. An array of polypeptides, said array of polypeptides comprising:

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

a plurality of different polypeptides bound to said first surface of said planar rigid support at a density exceeding 400 different polypeptides/cm², wherein each of said different polypeptides is attached to said first surface of said planar rigid support and has a different determinable sequence.

40. The array of claim 39, wherein said density exceeds 1000 different polypeptides occupying a total area of less than 1 cm² on said planar rigid support.

41. The array of claim 39, wherein said plurality of different polypeptides exceeds 1000 different polypeptides wherein each of said plurality of different polypeptides is attached to said first surface in a different positionally defined location of area greater than 100 square microns and has a different determinable sequence.