METHODS AND COMPOSITIONS FOR HYBRIDIZING NUCLEIC ACIDS

• US 20100151473A1
• Filed: 12/09/2009
• Published: 06/17/2010
• Est. Priority Date: 12/10/2008
• Status: Abandoned Application

• Current Assignee: Illumina Incorporated
• Original Assignee: Illumina Incorporated
• Inventors: Shen, Min-Jui Richard, Lenta, Ross Edward, Yeakley, Joanne M.
• Application Number:
  US12/634,531
• Publication Number:
  US 20100151473A1
• Time in Patent Office: Days
• US Class Current: 435/6
• CPC Class Codes: C12Q 1/6832 Enhancement of hybridisation reaction
• CPC Class Codes: C12Q 2527/125 Specific component of sample, medium or buffer

Abstract

Methods and compositions for hybridizing nucleic acids are disclosed herein. Also disclosed herein are methods and compositions to detect a nucleic acid provided to an array.

Claims

1. A nucleic acid hybridization method, said method comprising:

obtaining a buffered solution comprising a double-stranded nucleic acid and a divalent cation;

converting said double-stranded nucleic acid to a single-stranded nucleic acid in said buffered solution, wherein said converting comprises digesting said double-stranded nucleic acid with a nuclease; and

hybridizing said single-stranded nucleic acid to a capture probe in said buffered solution.

2. The method of claim 1, wherein said double-stranded nucleic acid comprises DNA.

3. The method of claim 1, wherein said nuclease is selected from the group consisting of exonuclease III, T7 exonuclease and lambda exonuclease.

4. The method of claim 1, wherein said nuclease is lambda exonuclease.

5. The method of claim 1, wherein said buffer lacks a concentration of monovalent cations sufficient to substantially inhibit the activity of said nuclease.

6. The method of claim 1, wherein said buffer lacks monovalent cations.

7. The method of claim 1, wherein said buffer lacks a concentration of phosphate ions sufficient to substantially inhibit the activity of said nuclease.

8. The method of claim 1, wherein said buffer lacks phosphate ions.

9. The method of claim 1, wherein said divalent cation is present in said buffered solution at a concentration sufficient to permit hybridization of said single-stranded nucleic acid complementary to said capture probe.

10. The method of claim 1, wherein said divalent cation is selected from the group consisting of Mg²⁺, Mn²⁺, Zn²⁺ and combinations thereof.

11. The method of claim 1, wherein said buffered solution further comprises a polyamine.

12. The method of claim 1, wherein the pH of the buffered solution is at least 7.5.

13. The method of claim 12, wherein said buffered solution comprises Tris buffer.

14. The method of claim 1, wherein said capture probe is associated with a solid support.

15. The method of claim 14, wherein said solid support is a planar surface.

16. The method of claim 14, wherein said solid support is a microsphere.

17. The method of claim 16, wherein said microsphere is porous.

18. The method of claim 14, wherein said solid support is a fiber optic bundle.

19. The method of claim 1, wherein said capture probe is one of a plurality of capture probes.

20. The method of claim 19, wherein said plurality of capture probes is distributed on the surface of a substrate.

21. The method of claim 20, wherein said plurality of capture probes is orderly distributed.

22. The method of claim 20, wherein said plurality of capture probes is randomly distributed.

23. The method of claim 1 further comprising extending the 3′

end of said capture probe by providing a polymerase enzyme.

24. A method for detecting the presence of a nucleic acid complementary to a capture probe, said method comprising:

obtaining a buffered solution comprising a double-stranded nucleic acid and a divalent cation;

converting said double-stranded nucleic acid to a single-stranded nucleic acid in said buffered solution, wherein said converting comprises digesting said double-stranded nucleic acid with a nuclease;

providing said single-stranded nucleic acid to a capture probe in said buffered solution; and

determining whether said single-stranded nucleic acid hybridizes to said capture probe, wherein hybridization of said single-stranded nucleic acid to said capture probe indicates the presence of a nucleic acid complementary to said capture probe.

25. A hybridization composition comprising:

a solid support comprising a capture probe; and

a buffered solution in fluid communication with said capture probe, said buffered solution comprising a double-stranded nucleic acid, a divalent cation, and an nuclease for converting said double-stranded nucleic acid to a single-stranded nucleic acid.

26. A hybridization composition comprising:

a solid support comprising a capture probe; and

a buffered solution in fluid communication with said capture probe, said buffered solution comprising a nuclease, a single-stranded nucleic acid and a divalent cation present at a concentration sufficient to permit hybridization between the single-stranded nucleic acid and the capture probe provided that the single-stranded nucleic acid has sufficient complementarity to hybridize with the capture probe.