Hybridization assays on oligonucleotide arrays

• US 6,045,996 A
• Filed: 05/16/1996
• Issued: 04/04/2000
• Est. Priority Date: 10/26/1993
• Status: Expired due to Term

• Current Assignee: Affymetrix, Inc. (Thermo Fisher Scientific Incorporated)
• Original Assignee: Affymetrix, Inc. (Thermo Fisher Scientific Incorporated)
• Inventors: Oval, Michelle, McGall, Glenn, Gingeras, Thomas R., Cronin, Maureen T., Trulson, Mark, Miyada, Charles Garrett, Robinson, Claire
• Primary Examiner(s): Marschel, Ardin H.
• Application Number:
  US08/648,709
• Time in Patent Office: 1,419 Days
• Field of Search: 422/50, 422/68.1, 435/6, 435/810, 436/501, 935/77, 935/78
• US Class Current: 506/4
• 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 2219/00432 Photolithographic masks
• CPC Class Codes: B01J 2219/00529 DNA chips
• 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 2219/00626 Covalent
• CPC Class Codes: B01J 2219/00659 Two-dimensional arrays
• CPC Class Codes: B01J 2219/00711 Light-directed synthesis
• CPC Class Codes: B01J 2219/00722 Nucleotides
• 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: 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: C12Q 1/6827 for detection of mutation or polymorphism
• CPC Class Codes: C12Q 1/6832 Enhancement of hybridisation reaction
• 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 2527/125 Specific component of sample, medium or buffer
• CPC Class Codes: C12Q 2527/127 the enzyme inhibitor or activator used
• CPC Class Codes: C12Q 2563/149 : Particles, e.g. beads
• CPC Class Codes: C40B 40/06 : Libraries containing nucleotides or polynucleotides, or derivatives thereof
• CPC Class Codes: C40B 60/14 : for creating libraries

Abstract

This invention provides methods of performing nucleic acid hybridization assays on high-density substrate-bound oligonucleotide arrays involving including in the hybridization mixture an isostabilizing agent, a denaturing agent or a renaturation accelerant.

Claims

1. A method for performing a hybridization assay between a target nucleic acid molecule and an oligonucleotide array, the array comprising a surface to which are covalently attached oligonucleotide probes with different, known sequences, at discrete, known locations, the array having a density of at least 500 said known locations of said different known sequences per square cm, the method comprising incubating the array with a hybridization mixture comprising the target and betaine and determining the identity of probes to which the target has hybridized.

2. The method of claim 1 wherein the mixture comprises betaine in a concentration of about 1 M to about 10 M.

3. The method of claim 2 wherein the mixture comprises betaine in a concentration of about 4 M to about 6 M.

4. The method of claim 1 further comprising the step, after incubating the array, of washing the array with a solution that comprises betaine.

5. The method of claim 1 wherein the target further comprises a detectable label.

6. The method of claim 5 wherein the label is a fluorescent label.

7. The method of claim 6 wherein the fluorescent label is fluorescein.

8. The method of claim 1 wherein the array has a density of at least ten thousand features per square cm.

9. The method of claim 8 wherein the array has a density of at least one hundred thousand features per square cm.

10. The method of claim 9 wherein the array has a density of at least one million features per square cm.