Molecular adaptors for dye conjugates

• US 9,777,320 B2
• Filed: 08/25/2011
• Issued: 10/03/2017
• Est. Priority Date: 08/25/2010
• Status: Active Grant

• Current Assignee: Pacific Bioscience of California Incorporated (L'Oreal SA)
• Original Assignee: Pacific Bioscience of California Incorporated (L'Oreal SA)
• Inventors: Yue, Stephen, Shen, Gene, Brogley, Louis, Sebo, Lubomir, Wegener, Jeffrey
• Primary Examiner(s): Rodriguez-Garcia, Valerie
• Application Number:
  US13/218,439
• Publication Number:
  US 20120058473A1
• Time in Patent Office: 2,231 Days
• CPC Class Codes: C07D 209/14 Radicals substituted by nitrogen atoms, not forming part of a nitro radical
• CPC Class Codes: C07D 401/14 containing three or more hetero rings
• CPC Class Codes: C07D 405/14 containing three or more hetero rings
• CPC Class Codes: C07H 19/10 with the saccharide radical esterified by phosphoric or polyphosphoric acids
• CPC Class Codes: C07K 5/0815 with the first amino acid being basic
• CPC Class Codes: C07K 5/1019 with the first amino acid being basic
• CPC Class Codes: C09B 23/0025 the substituent being bound through an oxygen atom
• CPC Class Codes: C09B 23/0033 the substituent being bound through a sulfur atom
• CPC Class Codes: C09B 23/0066 the polymethine chain being part of a carbocyclic ring,(e.g. benzene, naphtalene, cyclohexene, cyclobutenene-quadratic acid)
• CPC Class Codes: C09B 23/06 three >CH- groups, e.g. carbocyanines
• CPC Class Codes: C09B 23/083 five >CH- groups
• CPC Class Codes: C09B 23/105 two >CH- groups
• CPC Class Codes: C09B 23/164 containing one nitrogen atom
• CPC Class Codes: C12Q 1/6844 Nucleic acid amplification reactions
• CPC Class Codes: C12Q 1/6869 Methods for sequencing
• CPC Class Codes: C12Q 1/6876 Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
• CPC Class Codes: G01N 2333/9126 DNA-directed DNA polymerase (2.7.7.7)
• CPC Class Codes: G01N 33/52 Use of compounds or compositions for colorimetric, spectrophotometric or fluorometric investigation, e.g. use of reagent paper and including single- and multilayer analytical elements immunological elements G01N33/54386; involving labelled immunochemicals G01N33/58; for haemoglobin or occult blood G01N33/72
• CPC Class Codes: G01N 33/582 with fluorescent label

Abstract

In various embodiments, the present invention provides fluorescent dyes that are linked to another species through an adaptor moiety. In an exemplary embodiment, the dye is linked to a polyphosphate nucleic acid through an adaptor. An adaptor can be a component of a linker. These conjugates find use in single molecule DNA sequencing and other applications. In various embodiments, the dye moiety is a cyanine dye. Cyanine dyes that are highly charged, such as those including multiple sulfonate, alkylsulfonate, carboxylate and/or alkylcarboxylate moieties are examples of cyanine dyes of use in the compounds of the invention.

Claims

1. A fluorescent cyanine dye of Formula I:

{R¹-(L¹)_a-A}_y-X 

 

(I)

wherein

R¹ is a fluorescent cyanine dye moiety, of Formula Ia;

2. The fluorescent cyanine dye of claim 1, wherein A is selected from:

3. The fluorescent cyanine dye of claim 1, wherein A is selected from:

4. A method of monitoring an enzyme reaction, said method comprising:

(a) forming a reaction mixture by contacting said enzyme with the fluorescent cyanine dye according to claim 1, wherein said dye is a substrate for said enzyme under conditions sufficient for said enzyme and said dye to react; and

(b) monitoring fluorescence of said reaction mixture.

5. The method according to claim 4, wherein said enzyme is a DNA polymerase and said fluorescent cyanine dye comprises a nucleic acid moiety which is said substrate for said enzyme.

6. The method according to claim 5, wherein said reaction mixture further comprises a template nucleic acid that complexes with said DNA polymerase, and wherein said enzyme reaction is template directed DNA synthesis.

7. The method according to claim 6, wherein said enzyme reaction is a component of a single molecule DNA sequencing analysis.

8. The fluorescent cyanine dye according to claim 1, wherein R¹ is a fluorescent cyanine dye moiety selected from:

9. The fluorescent cyanine dye according to claim 1, wherein R¹ is a fluorescent cyanine dye moiety selected from:

10. A fluorescent cyanine dye of a formula selected from:

11. A fluorescent cyanine dye of Formula I:

{R¹-(L¹)_a-A}_y-X 

 

(I)

wherein

R¹ is a fluorescent cyanine dye moiety, of Formula Ia;