Engineered fluorescent dye labeled nucleotide analogs for DNA sequencing

• US 7,973,146 B2
• Filed: 03/26/2009
• Issued: 07/05/2011
• Est. Priority Date: 03/26/2008
• Status: Active Grant

• Current Assignee: Pacific Bioscience of California Incorporated (L'Oreal SA)
• Original Assignee: Pacific Bioscience of California Incorporated (L'Oreal SA)
• Inventors: Bibillo, Arkadusz, Shen, Gene, Peluso, Paul
• Primary Examiner(s): Riley; Jezia
• Application Number:
  US12/412,068
• Publication Number:
  US 20090246791A1
• Time in Patent Office: 831 Days
• Field of Search: 536/4.1, 536/26.6, 536/24.3, 536/24.33, 435/6, 435/91.1, 435/91.2
• US Class Current: 536/4.1
• CPC Class Codes: C07H 19/04 Heterocyclic radicals containing only nitrogen atoms as ring hetero atom

Abstract

Engineered nucleotide compositions, having polymerase interacting components that improve the interactivity of the polymerase and the nucleotide, particularly for nucleic acid sequencing applications. Compositions include the interactive polymerases along with the nucleotide analogs. Kits, methods and systems are provided for analysis of nucleic acid synthesis reactions.

Claims

1. A nucleotide composition, comprising the structure:

N—

Y—

F

wherein N is a nucleoside polyphosphate or analog thereof;

F is a detectable label moiety; and

Y comprises a polymerase interacting linkage complex comprising a polymerase interacting group positioned from 20 to 40 angstroms from a nucleoside portion of the nucleoside polyphosphate.

2. The nucleotide composition of claim 1, wherein the polymerase interacting linkage complex (Y) comprises the structure:

-L₁-P-L₂-

where L₁ is a first linker group, P is a polymerase interacting group, and L₂ is a second linker group.

3. The nucleotide composition of claim 1, wherein the polymerase interacting linkage complex (Y) comprises the structure:

4. The nucleotide composition of claim 3, wherein the polymerase interacting component comprises a hydrophobic group having one or more phenyl groups.

5. The nucleotide composition of claim 4, wherein the polymerase interacting component comprises a hydrophobic group comprising a phenyl group, a biphenyl group, a triphenyl group and a coumarin or coumarin derivative group.

6. The nucleotide composition of claim 2, comprising a structure selected from:

7. The nucleotide composition of claim 6, wherein R₁ through R₁₁ are independently selected from oxygen, hydroxyl, guanidinium, amine, ammonium, iminium, oxonium, carboxyl, phosphate, phosphonate, thio, sulfate, sulfonate, thioate, and phosphothioate groups.

8. The nucleotide composition of claim 6, wherein R₁₁ comprises a group selected from SO₃, PO₃, OH, or a double bonded O.

9. A nucleotide composition, comprising the structure:

N—

Y—

F

where N is a nucleoside polyphosphate group or analog thereof coupled through a phosphate group other than an alpha phosphate group;

L is a detectable labeling group; and

Y comprises a polymerase interacting linkage complex having a polymerase interacting component selected from phenyl, biphenyl, triphenyl and a coumarin derivative positioned from 20 to 40 angstroms from a nucleoside portion of the nucleoside polyphosphate.

10. The composition of claim 9, wherein N comprises a naturally occurring nucleoside coupled to a polyphosphate group.