Polymeric tandem dyes with linker groups
Compounds useful as fluorescent or colored dyes are disclosed. In some embodiments, 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 , M 1 , M 2 , m, and n are as defined herein. Methods associated with preparation and use of such compounds is also provided.
1. A compound having the following structure (III):
or a stereoisomer, salt, or tautomer thereof, wherein:
M 1 and M 2 are different chromophores selected such that M 1 and M 2 form a FRET pair;
L 1 is, at each occurrence, an optional linker, provided that at least one occurrence of L 1 is present and comprises oxygen;
L 2 and L 3 are, at each occurrence, independently an optional alkylene, alkenylene, alkynylene, heteroalkylene, heteroalkenylene, heteroalkynylene or heteroatomic linker;
L 4 is, at each occurrence, independently an alkylene or heteroalkylene linker;
L 8 is absent;
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, heteroalkyl, —OP(═R a )(R b )R c , Q, or a protected form thereof, or L′;
R 4 is, at each occurrence, independently OH, SH, O − , S − , OR d or SR d ;
R 5 is, at each occurrence, independently oxo, thioxo, or absent;
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 , OL′, SR d , alkyl, alkoxy, heteroalkyl, heteroalkoxy, alkylether, alkoxyalkylether, phosphate, thiophosphate, phosphoalkyl, thiophosphoalkyl, phosphoalkylether, or thiophosphoalkylether;
R d is a counter ion;
Q is, at each occurrence, independently a moiety comprising a reactive group, or protected form thereof, capable of forming a covalent bond with an analyte molecule, a targeting moiety, a solid support or a complementary reactive group Q′;
L′ is, at each occurrence, independently a linker comprising a covalent bond to Q, a linker comprising a covalent bond to a targeting moiety, a linker comprising a covalent bond to an analyte molecule, a linker comprising a covalent bond to a solid support, a linker comprising a covalent bond to a solid support residue, a linker comprising a covalent bond to a nucleoside or a linker comprising a covalent bond to a further compound of structure (III);
m is, at each occurrence, independently an integer of zero or greater; and
n is an integer of one or greater.
2. The compound of claim 1 , wherein the compound has the following structure (IIIA):
wherein:
L 4 is —(OCH 2 CH 2 ) z —;
z is, at each occurrence, independently an integer from 1 to 100; and
m is, at each occurrence, independently an integer from 0 to 6.
3. The compound of claim 2 , wherein z is, at each occurrence, independently an integer from 1 to 30.
4. The compound of claim 2 , wherein m is, at each occurrence, independently an integer from 2-4.
5. The compound of claim 2 , wherein the compound has the following structure (IIIB):
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.
6. The compound of claim 1 , wherein L 1 comprises one of the following structures:
wherein
a, b, and c are each independently an integer ranging from 1-6.
7. The compound of claim 1 , wherein R 4 is, at each occurrence, independently OH, O − or OR d ; R 5 is, at each occurrence, oxo; and R 1 is, at each occurrence, H.
8. 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.
9. The compound of claim 1 , wherein Q has one of the following structures:
wherein each X is independently a halogen.
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.
11. The compound of claim 1 , wherein R 2 or R 3 has one of the following structures:
12. The compound of claim 1 , wherein at least one combination of M 1 and M 2 is a FRET pair with a J-value greater than about 1×10 10 .
13. The compound of claim 1 , wherein M 1 and M 2 are, at each occurrence, independently selected from the group consisting of:
i) 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, and ter-naphthyl moiety;
ii) p-terphenyl, perylene, azobenzene, phenazine, phenanthroline, acridine, thioxanthrene, chrysene, rubrene, coronene, cyanine, perylene imide, perylene amide, and derivatives thereof,
iii) a coumarin dye, resorufin dye, dipyrrometheneboron difluoride dye, ruthenium bipyridyl dye, thiazole orange dye, polymethine, and N-aryl-1,8-naphthalimide dye; or
iv) a coumarin dye, boron-dipyrromethene, rhodamine, cyanine, pyrene, perylene, perylene monoimide, 6-FAM, 5-FAM, 6-FITC, 5-FITC, and derivatives thereof.
14. The compound of claim 1 , wherein M 1 and M 2 at each occurrence, independently have one of the following structures:
15. The compound of claim 1 , wherein the compound is selected from:
wherein:
Bs has the following structure:
FITC has the following structure:
AF555 has the following structure:
Cy3 has the following structure:
AF700 refers to Alexa Fluor 700 having a CAS Registry No. of 1246956-22-8;
AF680 has the following structure:
AF647 has the following structure:
AF350 has the following structure:
PB has the following structure:
AF594 has the following structure:
and
dT has the following structure:
wherein:
R is H or a direct bond.
16. A composition comprising the compound of claim 1 one or more analyte molecules.
17. The compound of claim 1 , wherein at least one occurrence of L 4 is heteroalkylene.
18. The compound of claim 17 , wherein the heteroalkylene comprises alkylene oxide.
19. The compound of claim 18 , wherein the heteroalkylene comprises ethylene oxide.