Methods of enhancing functional performance of nucleic acid arrays

• US 6,806,361 B1
• Filed: 03/15/2001
• Issued: 10/19/2004
• Est. Priority Date: 03/17/2000
• Status: Active Grant

• Current Assignee: Affymetrix, Inc. (Thermo Fisher Scientific Incorporated)
• Original Assignee: Affymetrix, Inc. (Thermo Fisher Scientific Incorporated)
• Inventors: McGall, Glenn, Trulson, Mark, Kajisa, Lisa
• Primary Examiner(s): FREDMAN, JEFFREY NORMAN
• Application Number:
  US09/811,143
• Time in Patent Office: 1,314 Days
• Field of Search: 536/25.3, 536/24.3, 536/23.1, 536/22.1, 435/6
• US Class Current: 536/25.3
• 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: C07H 21/04 with deoxyribosyl as saccharide radical

Abstract

Methods are provided for preparing nucleic acid arrays on a support. In these methods a plurality of nucleic acids are synthesized on the support and the synthesis steps are followed by drying steps in which the array is exposed to a dry atmosphere following the synthesis steps.

Claims

1. A method of reducing pitting of a solid support in the preparation of a nucleic acid array on said solid support, wherein said solid support is non-porous and substantially planar or comprises substantially planar regions, said method comprising:

a) attaching a plurality of nucleic acids to said support to form an array; and

b) drying said array by exposing to a dry atmosphere for a period of at least 30 seconds.

2. A method in accordance with claim 1, wherein said attaching comprises a stepwise coupling of nucleic acid monomers to a nucleic acid probe array wherein each of said probes are from 5 to 50 monomers in length and said drying comprises following at least a portion of said stepwise coupling steps with a step of exposing the array to a dry atmosphere for a period of at least 50 seconds.

3. A method in accordance with claim 2, wherein said portion is at least about 70% of said coupling steps.

4. A method in accordance with claim 2, wherein said portion is at least abut 85% of said coupling steps.

5. A method in accordance with claim 2, wherein said portion is at least about 95% of said coupling steps.

6. A method accordance with claim 1, wherein each nucleic acid occupies a separate known region of the support, said attaching step (a) further comprises:

(i) activating a region of the support;

(ii) attaching a nucleotide to a first region, said nucleotide having a masked reactive site liked to a protecting group;

(iii) repeatng steps (i) and (ii) on other regions of said support whereby each of said other regions has bound thereto another nucleotide comprising a masked reactive site link to a protecting group, wherein said another nucleotide may be the same or different from that used in step (ii);

(iv) removing the protecting group from one of the nucleotides bound to one of the regions of the support to provide a region bearing a nucleotide having an unmasked reactive site;

(v) binding an additional nucleotide to the nucleotide with an unmasked reactive site;

(vi) repeating steps (iv) and (v) on regions of the support until a desired plurality of nucleic acids is synthesized, each nucleic acid occupying separate known regions of the support;

wherein at least a portion of said attaching and said binding steps are followed by drying steps wherein said solid support is exposed to a dry atmosphere for a period of at least 50 seconds.

7. A method in accordance with claim 1, wherein said attaching step (a) comprises the sequential steps of:

(i) removing a photoremovable protecting group from at least a first area of a surface of a substrate, said surface comprising immobilized nucleotides on said surface, said nucleotides with a photoremovable protective group, without removing a photoremovable protecting group from at least a second area of said surface;

(ii) simultaneously contacting said first area and said second area of said surface with a first nucleotide to couple said first nucleotide to said immobilized nucleotides in said first area, and not in said second area, said first nucleotide capped with said photoremovable protective group;

(iii) removing a photoremovable protecting group from at least a part of said first area of said surface and at least a part of said second area;

(iv) simultaneously contacting said first area and said second area of said surface with a second nucleotide to couple said second nucleotide to said immobilized nucleotides in at least a part of said first area at least a part of said area;

(v) performing additional irradiating and nucleotide contacting and coupling steps so that a matrix array of at least 100 nucleic acids having different sequences is formed on said support;

wherein at least a portion of said contacting steps are followed by drying steps wherein said solid support is exposed to a dry atmosphere selected from the group consisting of dry air, nitrogen, argon and mixtures thereof for a period of at least 50 seconds.

8. A method in accordance with claim 7, wherein said portion is at least about 70% of said contacting steps.

9. A method in with claim 7, wherein said portion is at least about 85% of said contacting steps.

10. A method in accordance with claim 7, wherein said portion is at least about 95% of said contacting steps.

11. A method in accordance with claim 9, wherein said array comprises at least 10 different nucleic acids.

12. A method in accordance with claim 9, wherein said array comprises at least 100 different nucleic acids.

13. A method in accordance with claim 9, wherein said array at least 1000 different nucleic acids.

14. A method in accordance with claim 9, wherein said array comprises at least 10,000 different nucleic acids.

15. A method in accordance with claim 9, wherein said array comprises at least 100,000 different nucleic acids.

16. A method in accordance with claim 9, wherein each different nucleic acid is in a region having an area of less than about 1 cm².

17. A method in accordance with claim 9, wherein each different nucleic acid is in a region having an area of less than about 1 mm².