US 8,759,526 B2 · Dyes, composite dyes, and oligonucleotides or polynucleotides comprising such dyes—useful or detection or quantification of desirable target molecules

Abstract

The present invention provides dyes, reactive dyes and labeled reagents that may be used in the detection or quantification of desirable target molecules, such as proteins and nucleic acids. Dyes are provided that may be used free in solution where the binding of the dye to the target molecule provides signal generation. Dyes are also provided that comprise reactive groups that may be used to attach the dyes to probes that will bind to desirable target molecules. The novel dyes of the present invention have been modified by the addition of charged and polar groups to provide beneficial properties.

A dye having the structure:

wherein
at least one of R8 and R21 or R7 and R22 are taken together to form a five-membered or six-membered saturated or unsaturated heteroalkyl ring;
at least one of R1, R2, R3, R6, R7, R8R9, R21 or R22 is Q, wherein Q is selected from a sulfoxide (SOR13), a sulfone (SO2CR13R14R15), a sulfonamide (SO2NR13R14), a phosphonate (PO3═), a phosphonate monoester (PO2−ER13) a phosphonate dioster (POER13ER14), a thiophosphate (PSO3═), a thiophosphate monoester (PSO2−ER13) a thiophosphate diester (PSOER13ER14), a thiophosphonate (PSO2═), a thiophosphonate monoester (PSO−ER13) a thiophosphonate diester (PSER13ER14), a phosphonamide (PONR13R14NR19R20), a thioanalogue of a phosphonamide, (PSNR13R14NR19R20), a phosphoramide (PONR13R14NR15NR19R20), a thioanalogue of a phosphoramide (PSNR13R14NR15NR19R20), a phosphoramidite (PO2R19NR13R14), a thioanalogue of a phosphoramidite (POSR19R13R14), wherein E is O or S;
wherein Q is attached directly, or indirectly through a saturated or unsaturated, linear or branched, substituted or unsubstituted carbon linker arm, which may further comprise one or more heteroatoms selected from sulfur, oxygen and nitrogen, and wherein when Q is a sulfonamide, it does not comprise a terminal reactive group or a linker joining the dye to a target molecule;
R13, R14, R15, R19 and R20 are independently selected from hydrogen, halogen, an amino group, a saturated or unsaturated, linear or branched, substituted or unsubstituted alkyl group, or when taken together R13 and R14 form a five-membered or six-membered saturated or unsaturated heteroalkyl ring;
wherein the remaining R1, R2, R3, R5, R6, R7, R8, R9, R21 and R22 are independently selected from hydrogen, Z, a saturated or unsaturated, linear or branched, substituted or unsubstituted alkyl group, a saturated or unsaturated, linear or branched, substituted or unsubstituted alkyl group alkoxy group, or when taken together, R2 and R3, R6 and R7, and R8 and R9 form a five-membered or six-membered ring;
Z is selected from a carboxyl group (CO2−), a carbonate ester (COER13), a sulfonate (SO3−), a sulfonate ester (SO2ER13), a sulfoxide (SOR13), a sulfone (SO2CR13R14R15), a sulfonamide (SO2NR13R14), a phosphate (PO4═), a phosphate monoester (PO2−ER13), a phosphate diester (PO2ER13ER14), a phosphonate (PO3═) a phosphonate monoester (PSO2−ER13) a phosphonate diester (POER13ER14), a thiophosphate (PSO3═), a thiophosphate monoester (PSO2−ER13) a thiophosphate diester (PSOER13ER14), a thiophosphonate (PSO2═), a thiophosphonate monoester (PSO−ER13) a thiophosphonate diester (PSER13ER14), a phosphonamide (PONR13R14NR19R20), a thioanalogue of a phosphonamide (PSNR13R14NR19R20), a phosphoramide (PONR13R14NR15NR19R20), a thioanalogue of a phosphoramide (PSNR13R14NR15NR19R20), a phosphoramidite (PO2R19NR13R14) or a thioanalogue of a phosphoramidite (POSR19NR13R14), and E is O or S;
wherein Z is attached directly or indirectly through a saturated or unsaturated, linear or branched, substituted or unsubstituted carbon linker arm, which may further comprise one or more heteroatoms selected from sulfur, oxygen and nitrogen;
and wherein any of R1, R2, R3, R5, R6, R7, R8, R9, R21 or R22 may further comprise a heteroatom containing side chain wherein said side chain is joined to the R group by a linkage selected from an ether linkage (—OR25), a thioether linkage (—SR25), and an amine linkage (—NR25R26 or —N+R25R26R27), and
wherein R25, R26 and R27 are each independently selected from hydrogen, Z, a saturated or unsaturated, linear or branched, substituted or unsubstituted alkyl group, a saturated or unsaturated, linear or branched, substituted or unsubstituted alkoxy group, or when taken together, R25 and R26, or R26 and R27 form a saturated or unsaturated five-membered or six-membered alkyl or heteroalkyl ring, and wherein any of R25, R26 or R27 may further comprise a heteroatom containing side chain.

7 more elements

Dyes, composite dyes, and oligonucleotides or polynucleotides comprising such dyes—useful or detection or quantification of desirable target molecules

Life of this patent

Dyes, composite dyes, and oligonucleotides or polynucleotides comprising such dyes—useful or detection or quantification of desirable target molecules

Life of this patent

Abstract

Claims32