Methods of making nucleic acid or oligonucleotide arrays

• US 6,600,031 B1
• Filed: 04/21/1998
• Issued: 07/29/2003
• 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): ZITOMER, STEPHANIE W
• Application Number:
  US09/063,933
• Time in Patent Office: 1,925 Days
• Field of Search: 435/6, 435/91.1, 435/91.2, 536/23.1, 536/24.3, 536/25.4, 935/77, 935/78, 422/129, 422/130, 422/149
• 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 2219/00612 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 2219/00648 : 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 19/073 : with 2-deoxyribosyl as the saccharide radical
• CPC Class Codes: C07H 19/173 : with 2-deoxyribosyl as the saccharide radical
• CPC Class Codes: C07H 21/04 : with deoxyribosyl as saccharide radical
• 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: 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: G01N 21/6458 : Fluorescence microscopy fluorescence microscopes per se G02B21/0076 and G02B21/16
• 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 7/0045 : with organic non-macromolecular light-sensitive compounds not otherwise provided for, e.g. dissolution inhibitors
• 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: Y02P 20/55 : Design of synthesis routes, e.g. reducing the use of auxiliary or protecting groups
• 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

Abstract

A synthetic strategy for the creation of large scale chemical diversity. Solid-phase chemistry, photolabile protecting groups, and photolithography are used to achieve light-directed spatially-addressable parallel chemical synthesis. Binary masking techniques are utilized in one embodiment. A reactor system, photoremovable protective groups, and improved data collection and handling techniques are also disclosed. A technique for screening linker molecules is also provided.

Claims

1. A method of making an array of oligonucleotides, which comprises:

attaching, through the use of an automated device, a plurality of oligonucleotides to a impermeable surface of a support, the oligonucleotides having different predetermined sequences and being attached at different localized areas having a width of less than 100 microns on the surface of the support.

10. The method of claim 1 or claim 6 wherein the automated device employs photolithography to make said array.

2. A method for constructing an array of oligonucleotides of length s and composed of different nucleotides, which method comprises:

a) applying precursors for the different nucleotides to a plurality of different regions of a surface,

b) applying precursors for the different nucleotides to a plurality of different regions amongst the plurality of different regions defined in (a),

c) repeating the process until each of said regions contains oligonucleotides of lengths, each of said regions having a width of less than 100 microns.

3. The method as claimed in claim 2, wherein the regions are organized on the surface in rows and columns.

11. A method according to claim 3 wherein the automated device employs photolithography.

4. A method for generating an array of oligonucleotides on a support material comprising the steps of:

a) coupling a nucleotide precursor to a first set of localized areas on the support;

b) coupling a nucleotide precursor to a second set of localized areas on the support;

c) and continuing the sequence of coupling steps until the desired array has been generated,

the couplings being effected at each localized area either to the surface of the support or to a nucleotide coupled in a previous step, each of said localized areas having a width of less than 100 microns.

12. The method of claim 4 or claim 8 wherein the coupling is carried out through an automated device.

13. The method of claim 12 wherein the automated device employs photolithography.

5. A method of synthesizing up to n^l different types of nucleic acids or oligonucleotides, each oligonucleotide or nucleic acid being made up of l nucleotides selected from a different set of n different types of nucleotides, where n and l are greater than 1, said method comprising:

a) attaching a plurality of one type of nucleotide from said set to at least a portion of a substrate to form a first layer of said first type of nucleotide, said first layer having a surface made up of n portions, each of said portions having a width of less than 100 microns, said nucleotides having a protective material which prevents additional nucleotides from attaching to said first layer of monomers;

b) removing said protective material from a first portion of the surface of said first layer;

c) attaching a first selected type of nucleotide from said set of n nucleotides to the nucleotides exposed by removal of the protective material to form a second layer of said first selected type of nucleotide over those portions of the nucleotides of said first type exposed by removal of said protective material; and

d) repeating steps b) and c) up to n−

1 times.

14. The method of claim 5 or claim 7 wherein the attaching is carried out by means of an automated device.

15. The method of claim 14 wherein the automated device employs photolithography.

6. A method of making an array of oligonucleotides, which comprises:

attaching, through the use of an automated device, a plurality of oligonucleotides to an impermeable surface of a support, the oligonucleotides having different predetermined sequences and being attached at different known localized areas providing in excess of 400 oligonucleotides per cm² on the surface of the support.

7. A method for constructing an array of oligonucleotides of length s and composed of different nucleotides, which method comprises:

a) applying precursors for the different nucleotides to a plurality of different regions of a surface,

b) applying precursors for the different nucleotides to a plurality of different regions amongst the plurality of different regions defined in (a),

c) repeating the process until each of said regions contains oligonucleotides of length s so as to provide in excess of 400 nucleotides per cm².

8. A method for generating an array of oligonucleotides on a support material comprising the steps of:

a) coupling a nucleotide precursor to a first set of locations on the support;

b) coupling a nucleotide precursor to a second set of locations on the support;

c) and continuing the sequence of coupling steps until the desired array has been generated,

the couplings being effected at each location either to the surface of the support or to a nucleotide coupled in a previous step, so as to provide all array of at least 400 nucleic acids per cm².

9. A method of synthesizing up to n^l different types of nucleic acids or oligonucleotides, each oligonucleotides or nucleic acid being made up of l nucleotides selected from a different set of n different types of nucleotides, where n and l are greater than 1, said method comprising:

a) attaching a plurality of one type of nucleotide from said set to at least a portion of a substrate to form a first layer of said first type of nucleotide, said first layer having a surface made up of n portions, said nucleotides having a protective material which prevents additional nucleotides from attaching to said first layer of monomers;

b) removing said protective material from a first portion of the surface of said first layer;

c) attaching a first selected type of nucleotide from said set of n nucleotides to the nucleotides exposed by removal of the protective material to form a second layer of said first selected type of nucleotide over those portions of the nucleotides of said first type exposed by removal of said protective material; and

d) repeating steps b) and c) up to n−

1 times so as to provide an array of at least 400 nucleic acids per cm².