ENERGY TRANSFER DYES WITH ENHANCED FLUORESCENCE

• US 20100280246A1
• Filed: 12/28/2006
• Published: 11/04/2010
• Est. Priority Date: 05/03/1996
• Status: Abandoned Application

• Current Assignee: Applied Biosystems, Inc. (Thermo Fisher Scientific Incorporated)
• Original Assignee: Applera Corporation (Thermo Fisher Scientific Incorporated)
• Inventors: Lee, Linda G., Spurgeon, Sandra L., Rosenblum, Barnett
• Application Number:
  US11/617,650
• Publication Number:
  US 20100280246A1
• Time in Patent Office: Days
• US Class Current: 546/37
• CPC Class Codes: C07D 311/82 Xanthenes
• CPC Class Codes: C07D 311/90 with hydrocarbon radicals, substituted by amino radicals, directly attached in position 9
• CPC Class Codes: C07D 405/12 linked by a chain containing hetero atoms as chain links
• CPC Class Codes: C07D 405/14 containing three or more hetero rings
• 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
• CPC Class Codes: C09B 11/04 derived from triarylmethanes , i.e. central C-atom is substituted by amino, cyano, alkyl
• CPC Class Codes: C09B 11/24 Phthaleins containing amino groups ; Phthalanes; Fluoranes; Phthalides; Rhodamine dyes; Phthaleins having heterocyclic aryl rings; Lactone or lactame forms of triarylmethane dyes
• CPC Class Codes: C09B 23/083 five >CH- groups
• CPC Class Codes: C09B 69/00 Dyes not provided for by a single group of this subclass
• CPC Class Codes: C12Q 1/6818 involving interaction of two or more labels, e.g. resonant energy transfer
• CPC Class Codes: C12Q 1/6869 Methods for sequencing
• CPC Class Codes: C12Q 2535/101 Sanger sequencing method, i.e. oligonucleotide sequencing using primer elongation and dideoxynucleotides as chain terminators
• CPC Class Codes: C12Q 2563/107 fluorescence
• CPC Class Codes: C12Q 2565/1015 labels being on the same oligonucleotide

Abstract

Novel linkers for linking a donor dye to an acceptor dye in an energy transfer fluorescent dye are provided. These linkers facilitate the efficient transfer of energy between a donor and acceptor dye in an energy transfer dye. One of these linkers for linking a donor dye to an acceptor dye in an energy transfer fluorescent dye has the general structure R₂₁Z₁C(O)R₂₂R₂₈ where R₂₁ is a C_1-5 alkyl attached to the donor dye, C(O) is a carbonyl group, Z₁ is either NH, sulfur or oxygen, R₂₂ is a substituent which includes an alkene, diene, alkyne, a five and six membered ring having at least one unsaturated bond or a fused ring structure which is attached to the carbonyl carbon, and R₂₈ includes a functional group which attaches the linker to the acceptor dye.

Claims

1. An energy transfer dye comprising:

a xanthene donor dye capable of absorbing light at a first wavelength and emitting excitation energy in response thereto;

a 4,7-dichlororhodamine acceptor dye capable of absorbing the excitation energy emitted by the donor dye and fluorescing at a second wavelength in response thereto; and

a non-nucleosidic linker linking the xanthene donor dye to the 4,7-dichlororhodamine acceptor dye.

2. The energy transfer dye of claim 1 in which the non-nucleosidic linker links the xanthene donor dye to the 4′

-, 5- or 6-ring position of the 4,7-dichlororhodamine acceptor dye.

3. The energy transfer dye of claim 1 in which the linker has a backbone that is less than 9 atoms in length.

4. The energy transfer dye of claim 1 in which the linker comprises a functional group selected from an alkene, a diene, an alkyne, a five membered ring having at least one unsaturated bond, a six membered ring having at least one unsaturated bond and a fused ring structure.

5. The energy transfer dye of claim 1 in which the xanthene donor dye is a fluorescein dye.

6. The energy transfer dye of claim 5 which further comprises a linking group suitable for attaching the energy transfer dye to another substance.

7. The energy transfer dye of claim 6 in which the linking group is attached to the energy transfer dye via the 4′

-position of the 4,7-dichlororhodamine acceptor dye or the 4′

-, 5- or 6-position of the fluorescein donor dye.

8. The energy transfer dye of claim 1 in which the 4,7-dichlororhodamine acceptor dye comprises structure (I):

9. The energy transfer dye of claim 1 in which the xanthene donor dye comprises structure (II):

10. The energy transfer dye of claim 1 which comprises structure (III):

11. The energy transfer dye of claim 1 which comprises structure (IV):

12. The energy transfer dye of claim 10 or claim 11 in which X¹ is carboxylate.

13. The energy transfer dye of claim 12 in which R¹, R², R³, R⁴, R⁵, R⁶, R⁷, R⁸, R⁹, R¹⁰ and X³ are each hydrogen.

14. The energy transfer dye of claim 12 in which R¹, R³, R⁵, R⁷, R⁸, R¹⁰ and X³ are each hydrogen;

R² and R⁴ are each ethyl; and

R⁶ and R⁹ are each methyl.

15. The energy transfer dye of claim 12 in which R¹, R², R³ and R⁴ are each methyl;

and R⁵, R⁶, R⁷, R⁸, R⁹, R¹⁰ and X³ are each hydrogen.

16. The energy transfer dye of claim 12 in which R¹ and R⁵, R² and R⁶, R³ and R⁸ and R⁴ and R⁹ are taken together with the atoms to which they are bonded to form six-membered rings;

and R⁷, R¹⁰ and X³ are each hydrogen.

17. The energy transfer dye of claim 12 in which R¹, R³, R⁴, R⁵, R⁷, R⁸, R⁹, R¹⁰ and X³ are each hydrogen;

R² is ethyl; and

R⁶ is methyl.

18. The energy transfer dye of claim 12 in which R¹, R⁵, R⁷, R⁸, R⁹, R¹⁰ and X³ are each hydrogen;

R³, R⁴ and R⁶ are each methyl; and

R² is ethyl.

19. The energy transfer dye of claim 12 in which R⁵, R⁶, R⁷, R¹⁰ and X³ are each hydrogen;

R¹ and R² are each methyl; and

R³ and R⁸ and R⁴ and R⁹ are taken together with the atoms to which they are bonded to form six-membered rings.

20. The energy transfer dye of claim 12 in which R¹, R², R⁵, R⁶, R⁷, R¹⁰ and X³ are each hydrogen;

and R³ and R⁸ and R⁴ and R⁹ are taken together with the atoms to which they are bonded to form six-membered rings.

21. The energy transfer dye of claim 12 in which R¹, R⁵, R⁶, R⁷, R¹⁰ and X³ are each hydrogen;

R² is ethyl; and

R³ and R⁸ and R⁴ and R⁹ are taken together with the atoms to which they are bonded to form six-membered rings.

22. The energy transfer dye of claim 12 in which R¹, R², R⁵, R⁶, R⁷, R⁸, R⁹, R¹⁰ and X³ are each hydrogen;

and R³ and R⁴ are each methyl.