Labelled nucleotides

• US 7,427,673 B2
• Filed: 12/04/2002
• Issued: 09/23/2008
• Est. Priority Date: 12/04/2001
• Status: Expired due to Term

• Current Assignee: Illumina Cambridge Limited (Illumina Incorporated)
• Original Assignee: Illumina Cambridge Limited (Illumina Incorporated)
• Inventors: Balasubramanian, Shankar, Liu, Xiaohai, Barnes, Colin, Swerdlow, Harold
• Primary Examiner(s): Riley; Jezia
• Application Number:
  US10/497,594
• Publication Number:
  US 20060160075A1
• Time in Patent Office: 2,120 Days
• Field of Search: 536/23.1, 536/26.6, 435/6, 435/91.1
• US Class Current: 536/23.1
• CPC Class Codes: C07H 19/06 Pyrimidine radicals
• CPC Class Codes: C07H 19/10 with the saccharide radical esterified by phosphoric or polyphosphoric acids
• CPC Class Codes: C07H 19/16 Purine radicals
• CPC Class Codes: C07H 19/20 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: C12Q 1/6869 Methods for sequencing
• CPC Class Codes: C12Q 2525/186 incorporating a non-extendable or blocking moiety
• CPC Class Codes: C12Q 2525/197 incorporating a spacer/coupling moiety
• CPC Class Codes: C12Q 2535/113 Cycle sequencing

Abstract

Nucleosides and nucleotides are disclosed that are linked to detectable labels via a cleavable linker group.

Claims

1. A nucleotide or nucleoside molecule, having a purine, pyrimidine, or deazapurine base that is linked to a detectable label via a chemically cleavable linker, wherein the chemically cleavable linker comprises a protected hydroxyl functionality and chemically cleaving the protected hydroxyl functionality releases the detectable label from the purine, pyrimidine, or deazapurine base, wherein the nucleotide or nucleoside molecule further comprises a 3′

hydroxyl blocking group, and the protected hydroxyl functionality and the 3′

hydroxyl blocking group are cleavable under identical conditions.

2. The nucleotide or nucleoside molecule of claim 1, wherein chemical cleavage of the chemically cleavable linker produces a hydroxyl group attached to the purine, pyrimidine, or deazapurine base.

3. The nucleotide or nucleoside molecule of claim 1, wherein chemical cleavage of the chemically cleavable linker produces a hydroxyl group attached to the detectable label.

4. The nucleotide or nucleoside molecule of claim 1, having a ribose or deoxyribose sugar moiety.

5. The nucleotide or nucleoside molecule of claim 4, wherein the ribose or deoxyribose sugar comprises a protecting group attached via the 2′

or 3′

oxygen atom.

6. The nucleotide or nucleoside molecule of claim 1, that is a deoxyribonucleotide triphosphate.

7. The nucleotide or nucleoside molecule of claim 1, wherein the detectable label is a fluorophore.

8. The nucleotide or nucleoside molecule of claim 1, wherein the chemically cleavable linker comprises a moiety cleavable using palladium.

11. The nucleotide or nucleoside molecule of claim 1, wherein the chemically cleavable linker comprises a cleavable allylic system.

12. The nucleotide or nucleoside molecule of claim 1, wherein the nucleotide or nucleoside molecule is incorporated into a polynucleotide molecule.

13. The polynucleotide molecule of claim 12, wherein the polynucleotide molecule is immobilised on a solid support.

14. The polynucleotide molecule of claim 13, wherein the solid support is a bead or microsphere.

15. The polynucleotide molecule of claim 13, wherein the solid support is a glass slide.

16. A composition comprising at least one of the nucleotide or nucleo side molecules of claim 1.

17. A polynucleotide array comprising multiple immobilised polynucleotide sequences, each polynucleotide comprising a nucleotide or nucleoside of claim 1.

9. A kit comprising at least two nucleotide or nucleoside molecules, wherein each nucleotide or nucleoside molecule comprises a purine, pyrimidine, or deazapurine base that is linked to a detectable label via a chemically cleavable linker, wherein the chemically cleavable linker comprises a protected hydroxyl functionality and chemically cleaving the protected hydroxyl functionality releases the detectable label from the purine, pyrimidine, or deazapurine base, and the nucleotide or nucleoside molecule further comprises a 3′

hydroxyl blocking group, and the protected hydroxyl functionality and the 3′

hydroxyl blocking group are cleavable under identical conditions.

10. The kit of claim 9, further comprising an enzyme and buffers appropriate for the action of the enzyme.