Ultra bright dimeric or polymeric dyes with rigid spacing groups
Compounds useful as fluorescent or colored dyes are disclosed. The compounds have the following structure (I) or a stereoisomer, tautomer or salt thereof, wherein R 1 , R 2 , R 3 , R 4 , R 5 , L 1 , L 2 , L 3 , L 4 , L 5 , A, M, m and n are as defined herein. Methods associated with preparation and use of such compounds are also provided.
1. A compound having the following structure (I):
or a stereoisomer, salt or tautomer thereof, wherein:
A has, at each occurrence, independently one of the following structures:
wherein a 1 , a 2 and a 3 are, at each occurrence, independently a 5, 6 or 7-membered carbocyclic or heterocyclic ring and L 6 is a direct bond or a linker, wherein the wavy lines indicate linkages to -O-L 5 - and -L 4 -O- and wherein in the compound of structure (I), -O-L 5 - is covalently linked to a ring atom of the a 1 and -L 4 -O- is covalently linked to a ring atom of the a 3 ;
M is, at each occurrence, independently a fluorescent or colored moiety comprising two or more carbon-carbon double bonds and at least one degree of conjugation;
L 1 is at each occurrence, independently either: i) an optional alkylene, alkenylene, alkynylene, heteroalkylene, heteroalkenylene, heteroalkynylene or heteroatomic linker; or ii) a linker comprising a functional group capable of formation by reaction of two complementary reactive groups;
L 2 , L 3 , L 4 and L 5 are, at each occurrence, independently an optional alkylene, alkenylene, alkynylene, heteroalkylene, heteroalkenylene, heteroalkynylene or heteroatomic linker;
R 1 is, at each occurrence, independently H, alkyl or alkoxy;
R 2 and R 3 are each independently H, OH, SH, alkyl, alkoxy, alkylether, —OP(═R a )(R b )R c , Q, a linker comprising a covalent bond to Q, a linker comprising a covalent bond to an analyte molecule, a linker comprising a covalent bond to a solid support or a linker comprising a covalent bond to a further compound of structure (I), wherein: R a is O or S; R b is OH, SH, O − , S − , OR d or SR d ; R c is OH, SH, O − , S − , OR d , SR d , alkyl, alkoxy, alkylether, alkoxyalkylether, phosphate, thiophosphate, phosphoalkyl, thiophosphoalkyl, phosphoalkylether or thiophosphoalkylether; and R d is a counter ion;
Q is, at each occurrence, independently a moiety comprising a reactive group capable of forming a covalent bond with an analyte molecule, a solid support or a complementary reactive group Q′;
m is, at each occurrence, independently an integer of zero or greater, provided that at least one occurrence of m is an integer of one or greater; and
n is an integer of one or greater.
2. The compound of claim 1 , wherein A, at each occurrence, independently has one of the following structures:
3. The compound of claim 1 , wherein for at least one occurrence of L 1 , L 1 -M has the following structure:
wherein L 1a and L 1b are each independently optional linkers.
4. The compound of claim 1 , wherein for at least one occurrence of L 1 , L 1 -M has the following structure:
wherein L 1a and L 1b are each independently optional linkers.
5. The compound of claim 1 , wherein L 1 is at each occurrence, independently an optional alkylene or heteroalkylene linker.
6. The compound of claim 1 , wherein the compound has the following structure (IA):
wherein:
x 1 , x 2 , x 3 , x 4 , x 5 and x 6 are, at each occurrence, independently an integer from 0 to 6.
7. The compound of claim 1 , wherein R 1 is H.
8. The compound of claim 1 , wherein R 2 and R 3 are each independently OH or —OP(═R a )(R b )R c .
9. The compound of claim 1 , wherein one of R 2 or R 3 is OH or —OP(═R a )(R b )R c , and the other of R 2 or R 3 is Q or a linker comprising a covalent bond to Q.
10. The compound of claim 1 , wherein one of R 2 or R 3 is OH or —OP(═R a )(R b )R c , and the other of R 2 or R 3 is a linker comprising a covalent bond to an analyte molecule or a linker comprising a covalent bond to a solid support, wherein the analyte molecule is a nucleic acid or polymer thereof, an amino acid or polymer thereof, an enzyme, receptor, receptor ligand, antibody, glycoprotein, aptamer or prion and the solid support is a polymeric bead or nonpolymeric bead.
11. The compound of claim 1 , wherein M is, at each occurrence, independently a dimethylaminostilbene, quinacridone, fluorophenyl-dimethyl-BODIPY, his-fluorophenyl-BODIPY, acridine, terrylene, sexiphenyl, porphyrin, benzopyrene, (fluorophenyl-dimethyl-difluorobora-diaza-indacene)phenyl, (bis-fluorophenyl-difluorobora-diaza-indacene)phenyl, quaterphenyl, bi-benzothiazole, ter-benzothiazole, bi-naphthyl, bi-anthracyl, squaraine, squarylium, 9, 10-ethynylanthracene or ter-naphthyl moiety.
12. The compound of claim 1 , wherein M is, at each occurrence, independently p-terphenyl, perylene, azobenzene, phenazine, phenanthroline, acridine, thioxanthrene, chrysene, rubrene, coronene, cyanine, perylene imide, or perylene amide or derivative thereof.
13. The compound of claim 1 , wherein M is, at each occurrence, independently a coumarin dye, resorufin dye, dipyrrometheneboron difluoride dye, ruthenium bipyridyl dye, energy transfer dye, thiazole orange dye, polymethine or N-aryl-1,8-naphthalimide dye.
14. The compound of claim 1 , wherein M is, at each occurrence, independently pyrene, perylene, perylene monoimide or 6-FAM or derivative thereof.
15. The compound of claim 1 , wherein M, at each occurrence, independently has one of the following structures:
16. The compound of claim 1 , wherein the compound has one of the following structures:
wherein:
m is from 1 to 10;
y 1 and y 2 are each independently from 1 to 6;
F, E, and Y have the following structures:
17. A method for visually detecting an analyte molecule, the method comprising:
(a) admixing the compound of claim 1 , wherein R 2 or R 3 is Q or a linker comprising a covalent bond to Q, with the analyte molecule, wherein Q is the reactive group capable of forming a covalent bond with the analyte;
(b) forming a conjugate of the compound and the analyte molecule; and
(c) detecting the conjugate by its visible properties.