Coumarin dye and use thereof in sequencing

Abstract

The present invention relates to a coumarin dye and the use thereof in sequencing. Specifically, provided is a compound as shown in formula I or a stereoisomer, tautomer, crystal form, salt, ester or solvate thereof. The compound has the advantages of high brightness and high stability, and exhibits strong fluorescence under blue-light excitation. When used for gene sequencing after labeling dNTPs, the compound significantly improves the sequencing signal, resolution and throughput, thereby significantly improving the sequencing quality.

Claims

1. A compound represented by Formula I or a stereoisomer, tautomer, crystal form, salt, ester, or solvate thereof, wherein: R 1 selected from hydrogen, -COOH, R 1a R 1b R 1c R 1’a R 1’b R 1’c R 1d R 1e R 1f R 1g R 1h R 1i R 1j R 1k R 1l R 1m R 1n R 1o R 1p R 1q R 1r R 1s R 1t R 1u R 1v R 1w R 1x R 1y Each is independently selected from hydrogen, C1-C6 alkyl, carboxyl (-COOH), and carboxylate ion (-COO). - ), sulfonic acid group (-SO3H), sulfonate ion (-SO3) - ), halogen, halogenated C1-C6 alkyl group, wherein the C1-C6 alkyl group is optionally selected from carboxyl group (-COOH), carboxylate ion (-COO) - ), sulfonic acid group (-SO3H), sulfonate ion (-SO3) - The group replaced by ) R 2 , R 3 , R 4 each independently is selected from the group consisting of hydrogen, hydroxyl, cyano, nitro, carboxyl, thiol, halogen, C1-C6-alkyl, C1-C6-alkoxy, C1-C6-alkylthio, halogen-C1-C6-alkyl, C2-C6-alkenyl, C2-C6-alkynyl, C3-C6-cycloalkyl; R 5a , R 5b , R 6a , R 6b each independently is selected from the group consisting of hydrogen, hydroxyl, cyano, nitro, carboxyl, thiol, halogen, C1-C6-alkyl, C1-C6-alkoxy, C1-C6-alkylthio, halogen-C1-C6-alkyl, C2-C6-alkenyl, C2-C6-alkynyl, C3-C6-cycloalkyl; R 7 selected from -COOH, -SO3H; n is selected from 1, 2, 3, 4, 5, 6, preferably from 2, 3, 4, more preferably 3; X is selected from O, S, preferably O.

2. The compound of claim 1 or a stereoisomer, tautomer, crystal form, salt, ester, or solvate thereof, wherein, R 1 selected from hydrogen, -COOH, Preferably, R 1 selected from hydrogen, -COOH, More preferably, R 1 selected from hydrogen, -COOH, Most preferably, R 1 selected from hydrogen, -COOH, 3. The compound of any one of claims 1-2, or a stereoisomer, tautomer, crystal form, salt, ester, or solvate thereof, wherein, R 1a R 1b R 1c R 1d Each is independently selected from hydrogen, C1-C6 alkyl, carboxyl (-COOH), and carboxylate ion (-COO). - ), sulfonic acid group (-SO3H), sulfonate ion (-SO3) - ), halogen, halogenated C1-C6 alkyl group, wherein the C1-C6 alkyl group is optionally selected from carboxyl group (-COOH), carboxylate ion (-COO) - ), sulfonic acid group (-SO3H), sulfonate ion (-SO3) - The group replaced by ) Or, R 1a R 1b R 1c R 1d In, any one (such as R) 1b or R 1c ) or any two (such as R) 1b and R 1c Selected from hydrogen, C1-C6 alkyl, carboxyl (-COOH), and carboxylate ion (-COO) - ), sulfonic acid group (-SO3H), sulfonate ion (-SO3) - ), halogen, halogenated C1-C6 alkyl, wherein the C1-C6 alkyl group is optionally selected from carboxyl (-COOH), carboxylate ion (-COO) - ), sulfonic acid group (-SO3H), sulfonate ion (-SO3) - The group is replaced by hydrogen; Preferably, R 1a , R 1b , R 1c , R 1d are each independently selected from the group consisting of hydrogen, Ci-C6alkyl, carboxyl (-COOH), sulfonic acid group (-SO3H), halogen, halogenated Ci-C6alkyl, said Ci-C6alkyl being optionally substituted with a group selected from carboxyl (-COOH), sulfonic acid group (-SO3H); or preferably, R 1a , R 1b , R 1c , R 1d , any one (e.g. R 1b or R 1c ) or any two (e.g. R 1b and R 1c ) is selected from hydrogen, C1-C6alkyl, carboxyl (-COOH), sulfonic acid group (-SO3H), halogen, halogenated C1-C6alkyl, and said C1-C6alkyl is optionally substituted with a group selected from carboxyl (-COOH), sulfonic acid group (-SO3H), the remainder being hydrogen; More preferably, R 1a , R 1b , R 1c , R 1d each independently is selected from hydrogen, C1-C6alkyl, sulfonic acid group (-SO3H), carboxyl group (-COOH), halogen, halogenated C1-C6alkyl, said C1-C6alkyl being optionally substituted with a sulfonic acid group (-SO3H) or a carboxyl group (-COOH); or more preferably, R 1a , R 1b , R 1c , R 1d , any one (e.g., R 1b or R 1c ) or any two (e.g., R 1b and R 1c ) is selected from hydrogen, C1-C6alkyl, sulfonic acid group (-SO3H), carboxyl group (-COOH), halogen, halogenated C1-C6alkyl, and said C1-C6alkyl is optionally substituted with a sulfonic acid group (-SO3H) or a carboxyl group (-COOH), and the remainder is hydrogen; Further preferably, R 1a , R 1b , R 1c , R 1d are each independently selected from the group consisting of hydrogen, sulfo (-SO3H), carboxyl groups (-COOH), halo, halo-Ci-C6-alkyl; p is selected from 1, 2, 3, 4, 5, 5, preferably from 2, 3, 4; r is selected from 1, 2, 3, 4, 5, 8, preferably from 1, 2, 3; or further preferably, R 1a , R 1b , R 1c , R 1d , any one of which (e.g. R 1b or R 1c ) is selected from hydrogen, sulfo (-SO3H), carboxyl groups (-COOH), halogen, halogenated C1-C6alkyl, the rest are hydrogen, or, any two (such as R 1b and R 1c ) are selected from hydrogen, halogen, the rest are hydrogen; wherein, p is selected from 1, 2, 3, 4, 5, 6, preferably selected from 2, 3, 4; r is selected from 1, 2, 3, 4, 5, 6, preferably selected from 1, 2, 3; R 1a R 1b R 1c R 1d are each independently selected from the group consisting of hydrogen, a sulfonic acid group (-SO3H), carboxyl groups (-COOH), F, CF3; or more preferably, R 1a , R 1b , R 1c , R 1d , any one of which (e.g., R 1b or R 1c ) is selected from hydrogen, a sulfonic acid group (-SO3H), carboxyl groups (-COOH), - F, -CF3, the others being hydrogen, or, any two (such as R 1b and R 1c ) are selected from hydrogen, -F, the others being hydrogen; Most preferably, R 1a , R 1b , R 1c , R 1d are each independently selected from hydrogen, sulfonic acid group (-SO3H), carboxyl group (-COOH), -F, -CF3; or most preferably, R 1a , R 1b , R 1c , R 1d , any one (e.g. R 1b or R 1c ) is selected from hydrogen, sulfo (-SO3H), carboxyl (-COOH), -F, -CF3, and the remainder are hydrogen, or, any two (e.g. R 1b and R 1c ) are selected from hydrogen, -F, and the remainder are hydrogen; R 1e selected from hydrogen, C1-C6alkyl, carboxyl (-COOH), carboxylate ion (-COO - ), sulfonic acid group (-SO3H), sulfonate ion (-SO3 - ), said C1-C6alkyl being optionally substituted with a group selected from carboxyl (-COOH), carboxylate ion (-COO - ), sulfonic acid group (-SO3H), sulfonate ion (-SO3 - ). Preferably, R 1e is selected from hydrogen, C1-C6alkyl, carboxyl (-COOH), sulfonic acid group (-SO3H), said C1-C6alkyl being optionally substituted with a group selected from carboxyl (-COOH), sulfonic acid group (-SO3H); More preferably, R is selected from hydrogen, C1-C6alkyl, sulfo (-SO3H), said C1-C6alkyl being optionally substituted with sulfo (-SO3H); 1e selected from hydrogen, C1-C6alkyl, sulfo (-SO3H), said C1-C6alkyl being optionally substituted with sulfo (-SO3H); Further preferably, R 1e selected from hydrogen, sulfo group (-SO3H), p is selected from 1, 2, 3, 4, 5, 7, preferably from 2, 3, 4; More preferably, R 1e selected from hydrogen, sulfo group (-SO3H), Most preferably, R 1e selected from hydrogen, R 1f selected from the group consisting of C1-C6 alkyl, carboxylate ion (-COO - ), sulfonate ion (-SO3 - ), said C1-C6 alkyl being substituted with a group selected from the group consisting of carboxylate ion (-COO - ), sulfonate ion (-SO3 - ). Preferably, R 1f selected from the group consisting of Ci-C6-alkyl, sulfonate ion (-SO3 - ), said Ci-C6-alkyl being substituted by a sulfonate ion (-SO3 - ), or preferably, R 1f selected from sulfonate ions (-SO3 - ), carboxylate ion (-COO - ), q is selected from 1, 2, 3, 4, 5, 9, preferably from 2, 3, 4; t is selected from 1, 2, 3, 4, 5, 10, preferably from 1, 2, 3; More preferably, R 1f selected from sulfonate ions (-SO3 - ), carboxylate ion (-COO - ), Further preferably, R 1f is selected from the group consisting of sulfonate ions (-SO3 - ), Most preferably, R 1f is R 1g , R 1h each independently is selected from the group consisting of hydrogen, Ci-C6alkyl, carboxyl (-COOH), carboxylate ion (-COO - ), sulfonic acid group (-SO3H), sulfonate ion (-SO3 - ), said Ci-C6alkyl being optionally substituted with a group selected from carboxyl (-COOH), carboxylate ion (-COO - ), sulfonic acid group (-SO3H), sulfonate ion (-SO3 - ). Preferably, R 1g , R 1h each independently is selected from hydrogen, C1-C6alkyl, carboxyl (-COOH), sulfonic acid group (-SO3H), said C1-C6alkyl being optionally substituted with a group selected from carboxyl (-COOH), sulfonic acid group (-SO3H); More preferably, R 1g , R 1h each independently is selected from the group consisting of hydrogen, C1-C6alkyl, sulfonic acid group (-SO3H), said C1-C6alkyl being optionally substituted with a sulfonic acid group (-SO3H); Further preferably, R 1g , R 1h are each independently selected from the group consisting of hydrogen, a sulfonic acid group (-SO3H), p is selected from 1, 2, 3, 4, 6, preferably from 2, 3, 4; Most preferably, R 1g , R 1h are each independently selected from hydrogen, a sulfonic acid group (-SO3H), R 1i selected from the group consisting of C1-C6 alkyl, carboxylate ion (-COO - ), sulfonate ion (-SO3 - ), said C1-C6 alkyl being substituted with a group selected from the group consisting of carboxylate ion (-COO - ), sulfonate ion (-SO3 - ); Preferably, R 1i selected from sulfonate ions (-SO3 - ), carboxylate ion (-COO - ), q is selected from 1,2, 3, 4, 5, 6, preferably from 2,3, 4; t is selected from 1, 2, 3, 5, 6, preferably from 1, 2, 3; More preferably, R 1i selected from sulfonate ions (-SO3 - ), carboxylate ion (-COO - ), Most preferably, R 1i is R 1j , R 1k each independently is selected from the group consisting of C1-C6-alkyl, carboxylate ion (-COO - ), sulfonate ion (-SO3 - ), said C1-C6-alkyl being substituted by a group selected from the group consisting of carboxylate ion (-COO - ), sulfonate ion (-SO3 - ); Preferably, R 1j , R 1k are each independently selected from the group consisting of sulfonate ions (-SO3 - ), carboxylate ion (-COO - ), q is selected from 1, 2, 3, 4, 5, 6, preferably from 2, 3, 4; t is selected from 1,2, 3, 4, 5, 6, preferably from 1, 2, 3; Most preferably, R 1j , R 1k are each independently selected from the group consisting of sulfonate ions (-SO3 - ), carboxylate ion (-COO - ), R 1’a , R 1’b , R 1’c each independently is selected from the group consisting of hydrogen, Ci-C6alkyl, carboxyl (-COOH), carboxylate ion (-COO - ), sulfonic acid group (-SO3H), sulfonate ion (-SO3 - ), said Ci-C6alkyl being optionally substituted with a group selected from carboxyl (-COOH), carboxylate ion (-COO - ), sulfonic acid group (-SO3H), sulfonate ion (-SO3 - ). Preferably, R 1’a , R 1’b , R 1’c are each independently selected from the group consisting of hydrogen, C1-C6alkyl, carboxyl (-COOH), sulfonic acid group (-SO3H), said C1-C6alkyl being optionally substituted with a group selected from carboxyl (-COOH), sulfonic acid group (-SO3H); More preferably, R 1’a , R 1’b , R 1’c are each independently selected from the group consisting of hydrogen, sulfo (-SO3H), carboxyl groups (-COOH), p is selected from 1, 3, 4, 5, 6, preferably from 2, 3, 4; r is selected from 1, 2, 3, 4,5, 6, preferably from 1, 2, 3; Further preferably, R 1’a , R 1’b , R 1’c are each independently selected from the group consisting of hydrogen, sulfo (-SO3H), carboxyl groups (-COOH), R 1’a R 1’b R 1’c are each independently selected from the group consisting of hydrogen, Most preferably, R 1’a , R 1’b , R 1’c are each hydrogen; R 1l , R 1m each independently is selected from the group consisting of hydrogen, Ci-C6alkyl, carboxyl (-COOH), carboxylate ion (-COO - ), sulfonic acid group (-SO3H), sulfonate ion (-SO3 - ), said Ci-C6alkyl being optionally substituted with a group selected from carboxyl (-COOH), carboxylate ion (-COO - ), sulfonic acid group (-SO3H), sulfonate ion (-SO3 - ). Preferably, R 1l , R 1m each independently is selected from the group consisting of hydrogen, C1-C6alkyl, carboxyl (-COOH), sulfonic acid group (-SO3H), said C1-C6alkyl being optionally substituted with a group selected from carboxyl (-COOH), sulfonic acid group (-SO3H); More preferably, R 1l , R 1m are each independently selected from the group consisting of hydrogen, a sulfonic acid group (-SO3H), carboxyl groups (-COOH), p is selected from 1,2, 3, 4, 5, 6 preferably from 2, 3, 4; r is selected from 1, 3, 4, 5, 6, preferably 1, 2, 3; Further preferably, R 1l , R 1m are each independently selected from the group consisting of hydrogen, a sulfonic acid group (-SO3H), carboxyl groups (-COOH), Still more preferably, R 1l , R 1m are each independently selected from hydrogen, Still further preferably, R 1l , R 1m are each independently selected from hydrogen, Most preferably, R 1l , R 1m , R 1l is selected from hydrogen, another is hydrogen; R 1n , R 1o each independently is selected from the group consisting of hydrogen, Ci-C6alkyl, carboxyl (-COOH), carboxylate ion (-COO - ), sulfonic acid group (-SO3H), sulfonate ion (-SO3 - ), said Ci-C6alkyl being optionally substituted with a group selected from carboxyl (-COOH), carboxylate ion (-COO - ), sulfonic acid group (-SO3H), sulfonate ion (-SO3 - ). Preferably, R 1n , R 1o each independently is selected from hydrogen, C1-C6alkyl, carboxyl (-COOH), sulfonic acid group (-SO3H), said C1-C6alkyl being optionally substituted with a group selected from carboxyl (-COOH), sulfonic acid group (-SO3H); More preferably, R 1n , R 1o are each independently selected from the group consisting of hydrogen, a sulfonic acid group (-SO3H), carboxyl groups (-COOH), p is selected from 1, 2, 3, 4, 5, 7, preferably from 2, 3, 4; r is selected from 1,2, 3, 4, 5, 6; Further preferably, R 1n , R 1o are each independently selected from the group consisting of hydrogen, a sulfonic acid group (-SO3H), carboxyl groups (-COOH), R is more preferably selected from hydrogen, 1n R is more preferably selected from hydrogen, 1o each independently selected from hydrogen, Most preferably, R 1n , R 1o are each hydrogen; R 1p , R 1q , R 1r each independently is selected from the group consisting of hydrogen, Ci-C6alkyl, carboxyl (-COOH), carboxylate ion (-COO - ), sulfonic acid group (-SO3H), sulfonate ion (-SO3 - ), said Ci-C6alkyl being optionally substituted with a group selected from carboxyl (-COOH), carboxylate ion (-COO - ), sulfonic acid group (-SO3H), sulfonate ion (-SO3 - ). Preferably, R 1p , R 1q , R 1r each independently is selected from the group consisting of hydrogen, C1-C6alkyl, carboxyl (-COOH), sulfonic acid group (-SO3H), said C1-C6alkyl being optionally substituted with a group selected from carboxyl (-COOH), sulfonic acid group (-SO3H); More preferably, R 1p , R 1q , R 1r are each independently selected from the group consisting of hydrogen, sulfo (-SO3H), carboxyl groups (-COOH), p is selected from 1, 2, 3, 4, 5 6, preferably from 2, 3, 4; r is selected from 1,3, 4, 5, 6, preferably 1, 2, 5; Further preferably, R 1p , R 1q , R 1r are each independently selected from the group consisting of hydrogen, sulfo (-SO3H), carboxyl groups (-COOH), R 1p , R 1q , R 1r are each independently selected from the group consisting of hydrogen, Most preferably, R 1p , R 1q , R 1r are each hydrogen; R 1s , R 1t , R 1u , R 1v are each independently selected from the group consisting of hydrogen, Ci-C6alkyl, carboxyl (-COOH), carboxylate ion (-COO - ), sulfonic acid group (-SO3H), sulfonate ion (-SO3 - ), said Ci-C6alkyl being optionally substituted with a group selected from carboxyl (-COOH), carboxylate ion (-COO - ), sulfonic acid group (-SO3H), sulfonate ion (-SO3 - ); R 1s , R 1t , R 1u , R 1v are each independently selected from the group consisting of hydrogen, C1-C6alkyl, carboxyl (-COOH), sulfonic acid group (-SO3H), said C1-C6alkyl being optionally substituted with a group selected from carboxyl (-COOH), sulfonic acid group (-SO3H); More preferably, R 1s , R 1t , R 1u , R 1v are each independently selected from the group consisting of hydrogen, sulfo (-SO3H), carboxyl groups (-COOH), p is selected from 1, 2, 3, 4, 5. 6, preferably from 2, 3, 4; r is selected from 2, 3, 4, 5, 6, preferably 1, 2,3; Further preferably, R 1s , R 1t , R 1u , R 1v are each independently selected from the group consisting of hydrogen, sulfo (-SO3H), carboxyl groups (-COOH), R 1s , R 1t , R 1u , R 1v are each independently selected from the group consisting of hydrogen, Most preferably, R 1s , R 1t , R 1u , R 1v are each hydrogen; R 1w , R 1x , R 1y are each independently selected from the group consisting of hydrogen, Ci-C6alkyl, carboxyl (-COOH), carboxylate ion (-COO - ), sulfonic acid group (-SO3H), sulfonate ion (-SO3 - ), said Ci-C6alkyl being optionally substituted with a group selected from carboxyl (-COOH), carboxylate ion (-COO - ), sulfonic acid group (-SO3H), sulfonate ion (-SO3 - ). Preferably, R 1w , R 1x , R 1y each independently is selected from the group consisting of hydrogen, C1-C6alkyl, carboxyl (-COOH), sulfonic acid group (-SO3H), said C1-C6alkyl being optionally substituted with a group selected from carboxyl (-COOH), sulfonic acid group (-SO3H); More preferably, R 1w , R 1x , R 1y are each independently selected from the group consisting of hydrogen, sulfo (-SO3H), carboxyl groups (-COOH), p is selected from 1, 2, 3, 4, 7, preferably from 2, 3, 4; r is selected from 2,3, 4, 5, 6, preferably 1, 2, 4; Further preferably, R 1w , R 1x , R 1y are each independently selected from the group consisting of hydrogen, sulfo (-SO3H), carboxyl groups (-COOH), R 1w R 1x R 1y are each independently selected from the group consisting of hydrogen, Most preferably, R 1w , R 1x , R 1y are each hydrogen.

4. The compound according to any one of claims 1-3, or a stereoisomer, tautomer, crystal form, salt, ester, or solvate thereof, wherein, R 2 , R 3 , R 4 each independently is selected from the group consisting of hydrogen, hydroxyl, cyano, nitro, carboxyl, thiol, halogen, Ci-C6-alkyl, halogenated Ci-C6-alkyl; R 2 , R 3 , R 4 are each independently selected from the group consisting of hydrogen, Ci-C6alkyl, halogenated Ci-C6alkyl; or preferably, R 2 is selected from hydrogen, C1-C6alkyl, haloC1-C6alkyl, R 3 , R 4 is hydrogen; More preferably, R 2 , R 3 , R 4 are each independently selected from hydrogen, methyl, trifluoromethyl; Most preferably, R 2 is selected from hydrogen, methyl, trifluoromethyl, R 3 , R 4 is hydrogen.

5. The compound of any one of claims 1-4, or a stereoisomer, tautomer, crystal form, salt, ester, or solvate thereof, wherein, R 5a , R 5b , wherein any one is selected from the group consisting of hydrogen, hydroxy, cyano, nitro, carboxy, mercapto, halogen, C1-C6-alkyl, C1-C6-alkoxy, C1-C6-alkylthio, halogen-C1-C6-alkyl, C2-C6-alkenyl, C2-C6-alkynyl, C3-C6-cycloalkyl, and the other is hydrogen; R 5a , R 5b each independently is selected from the group consisting of hydrogen, hydroxyl, cyano, nitro, carboxyl, thiol, halogen, Ci-C6-alkyl; Or preferably, R 5a R 5b In this group, one of the radicals is selected from hydrogen, hydroxyl, cyano, nitro, carboxyl, mercapto, halogen, C1-C6 alkyl, and the other is hydrogen; More preferably, R 5a , R 5b each independently is selected from hydrogen, Ci-C6alkyl; or more preferably, R 5a , R 5b , wherein any one is selected from hydrogen, C1-C6alkyl, and the other is hydrogen; Most preferably, R 5a , R 5b are each independently selected from hydrogen, methyl; or most preferably, R 5a , R 5b , wherein either one is selected from hydrogen, methyl, the other is hydrogen; R 6a , R 6b each independently is selected from the group consisting of hydrogen, hydroxyl, cyano, nitro, carboxyl, thiol, halogen, C1-C6-alkyl, C1-C6-alkoxy, C1-C6-alkylthio, halogen-C1-C6-alkyl, C2-C6-alkenyl, C2-C6-alkynyl, C3-C6-cycloalkyl; Preferably, R 6a , R 6b each independently is selected from the group consisting of hydrogen, hydroxyl, cyano, nitro, carboxyl, thiol, halogen, Ci-C6-alkyl; More preferably, R 6a , R 6b are each independently selected from hydrogen, Ci-C6alkyl; Most preferably, R 6a , R 6b are each independently selected from hydrogen, methyl.

6. The compound of any one of claims 1-5, or a stereoisomer, tautomer, crystal form, salt, ester, or solvate thereof, wherein, The compound has a structural formula shown in formula I-1, wherein: R 1-1 selected from Preferably, R 1-1 selected from More preferably, R 1-1 selected from Further preferably, R 1-1 selected from Most preferably, R 1-1 selected from R 1a , R 1b , R 1c , R 1’a , R 1’b , R 1’c , R 1d , R 1e , R 1f , R 1g , R 1h , R 1i , R 1j , R 1k , R 2 , R 3 , R 4 , R 5a , R 5b , R 6a , R 6b , R 7 , n, X are each independently as defined in any one of claims 1 to 5.

7. The compound of any one of claims 1-5, or a stereoisomer, tautomer, crystal form, salt, ester, or solvate thereof, wherein, The compound has a structural formula shown in formula I-2, wherein: R 1-2 selected from Preferably, R 1-2 selected from More preferably, R 1-2 selected from Further preferably, R 1-2 selected from Most preferably, R 1-2 selected from R 1e , R 1f , R 1i , R 1j , R 1k , R 1l , R 1m , R 1n , R 1o , R 1p , R 1q , R 1r , R 1s , R 1t , R 1u , R 1v , R 1w , R 1x , R 1y , R 2 , R 3 , R 4 , R 5a , R 5b , R 6a , R 6b , R 7 , n, X are each independently as defined in any one of claims 1 to 5.

8. The compound of any one of claims 1-7, or a stereoisomer, tautomer, crystal form, salt, ester, or solvate thereof, wherein, The compound is selected from:

9. An intermediate having the structural formula of Formula I-i, ###0007### wherein: R 2 , R 3 , R 4 , R 5a , R 5b , R 6a , R 6b , n are each independently as defined in any one of claims 1 to 5; R 8 selected from -SO3H, -C(=O)-O-Ci-C6-alkyl, preferably from -SO3H, 10. The intermediate of claim 9 selected from the group consisting of:

11. A modified nucleoside, nucleotide or oligonucleotide comprising a nucleoside, nucleotide or oligonucleotide, and a detectable label; said nucleoside, nucleotide or oligonucleotide is linked to said detectable label, said detectable label is a compound according to any one of claims 1 to 8, or a stereoisomer, a tautomer, a crystal form, a salt, an ester or a solvate thereof; Preferably, said nucleoside, nucleotide or oligonucleotide is linked to the carboxyl group (-COOH) or the sulfonic acid group (-SO3H) of said detectable label; Preferably, the nucleoside, nucleotide or oligonucleotide comprises a base, a deaza base or a tautomer thereof, which is linked to the carboxyl (-COOH) or sulfonic acid group (-SO3H) of the detectable label; preferably, the base is selected from the group consisting of adenine, thymine, uracil, cytosine, guanine; preferably, the deaza base is selected from the group consisting of 7-deazaadenine, 7-deazaguanine; preferably, the nucleoside, nucleotide or oligonucleotide comprises The the 1st position of the nucleoside, nucleotide or oligonucleotide is linked to a carboxyl (-COOH) or sulfonic acid group (-SO3H) of the detectable label; preferably, the nucleoside, nucleotide or oligonucleotide further comprises a ribose or deoxyribose sugar, the the 2ndposition of is linked to the 1 '-carbon atom of said ribose or deoxyribose sugar; Preferably, said modified nucleoside, nucleotide or oligonucleotide further comprises a linker, said nucleoside, nucleotide or oligonucleotide is linked to said detectable label through the linker, said detectable label is a compound according to any one of claims 1 to 8 or a stereoisomer, a tautomer, a crystal form, a salt, an ether or a solvate thereof; Preferably, said nucleoside, nucleotide or olignucleotide is linked to the carboxyl group (-COOH) or the sulfonic acid (-SO3H) of said detectable label through the linker; Preferably, said nucleoside, nucleotide or olignucleotide is linked through the linker to the carboxyl group (-COOH) or the sulfonic acid (-SO3H). Preferably, the nucleoside, nucleotide or oligonucleotide comprises a base, a deaza base or a tautomer thereof linked to the carboxyl (-COOH) or sulfonic acid group (-SO3H) of the detectable label via a linker; preferably, the base is selected from the group consisting of adenine, thymine, uracil, cytosine, guanine; preferably, the deaza base is selected from the group consisting of 7-deazaadenine, 7-deazaguanine; preferably, the deaza base is selected from the group consisting of 7-deazaadenine, 7-deazaguanine; preferably, the nucleoside, nucleotide or oligonucleotide comprises The to the linker at position 1 of the nucleoside, nucleotide, or oligonucleotide; preferably, the nucleoside, nucleotide, or oligonucleotide further comprises a ribose or deoxyribose sugar, the is linked to the 1'-carbon atom of the ribose or deoxyribose sugar; Preferably, the nucleoside, nucleotide or oligonucleotide further comprises a reversible blocking group; preferably, the reversible blocking group is linked to the 3'-OH of the ribose or deoxyribose sugar comprised by the nucleoside, nucleotide or oligonucleotide.

12. The modified nucleoside, nucleotide, or oligonucleotide of claim 11, wherein, The modified nucleotides have a structure selected from the group consisting of: wherein T is -L-N, L is a linker, and N is a nucleotide; Preferably, the linker is Preferably, the nucleotide is selected from the group consisting of: wherein R 0 is a reversible blocking group; Preferably, the reversible blocking group is 13. The modified nucleoside, nucleotide, or oligonucleotide of claim 11 or 12, wherein, The modified nucleotides have a structure selected from the group consisting of:

14. A kit comprising a first nucleotide linked to a first detectable label, the first detectable label being a compound of any one of claims 1-8, or a stereoisomer, tautomer, crystal form, salt, ester, or solvate thereof; Preferably, the kit further comprises a first nucleotide linked to a second detectable label, the second detectable label being a compound different from the first detectable label, or a stereoisomer, tautomer, crystal form, salt, ester, solvate thereof; Preferably, the kit further comprises a second nucleotide linked to a first detectable label, the first detectable label being a compound of claims 1-8, or a stereoisomer, tautomer, crystal form salt, ester, or solvate thereof; Preferably, the kit further comprises third nucleotide linked to a second detectable label, the second detectable label being a compound different than the first detectable label, or a stereoisomer, tautomer, crystal form, a salt, an ester, or a solvate thereof; Preferably, the kit further comprises a fourth nucleotide not linked to a detectable label; Preferably, the first nucleotide linked to the first detectable label and the second nucleotide linked to the first detectable label are excitable by a first light source wavelength, the first nucleotide linked to the second detectable label and the third nucleotide linked to the second detectable label are excitable by a second light source wavelength, and the fourth nucleotide not linked to a detectable label is not excitable by the first light source wavelength and the second light source wavelength; Preferably, the kit further comprises a polymerase and one or more buffer solutions for performing a nucleotide polymerization reaction.

15. A method of determining the sequence of a target polynucleotide, comprising incorporating the modified nucleotide of any one of claims 11-13 into the complementary sequence of the target polynucleotide; Preferably, the method further comprises detecting the detectable label comprised by the modified nucleotide; Preferably, the method of determining the sequence of a target polynucleotide is performed on an automated sequencing instrument, and wherein the automated sequencing instrument comprises an excitation light source; preferably, the automated sequencing instrument comprises two excitation light sources operating at different wavelengths.

16. The method of claim 15, wherein, The method comprises: (A) incorporating at least one modified nucleotide of any one of claims 11-13 into the complementary sequence of the target nucleotide; and (B) determining the type of incorporated nucleotide by detecting the fluorescent signal of the detectable label comprised by the modified nucleotide.

17. The method of claim 15 or 16, comprising: (a) providing a plurality of different nucleotides, wherein at least one nucleotide is a modified nucleotide of any one of claims 11-13; (b) incorporating the plurality of different nucleotides into a complementary sequence of the target polynucleotide, wherein the plurality of different nucleotides are distinguishable from each other upon detection; (c) detecting the nucleotides of (b) to determine the type of nucleotide incorporated; (d) removing the detectable label and the reversible blocking group from the nucleotides of (b); and (e) optionally repeating steps (a)-(d) one or more times; thereby determining the sequence of the target polynucleotide; Preferably, the detectable label and the reversible blocking group are removed using a cleaving reagent. Preferably, the cleaving reagent that removes the detectable label and the reagent that removes the reversible blocking group are the same. Preferably, the removal of the detectable label and the removal of the reversible blocking group are performed simultaneously.

18. The method of any one of claims 15-17, comprising the steps of: (1) providing a first nucleotide, a second nucleotide, a third nucleotide, and a fourth nucleotide, at least one of the four nucleotides being a modified nucleotide of any one of claims 11-13; (2) contacting the four nucleotides with a target polynucleotide; removing the nucleotides that are not incorporated into the growing nucleic acid strand; detecting the nucleotides that are incorporated into the growing nucleic acid strand; removing the reversible blocking group and the detectable label contained in the nucleotides that are incorporated into the growing nucleic acid strand; Optionally, further comprising (3) repeating (1)-(2) one or more times.

19. The method of any one of claims 15-18, comprising the steps of: (a') providing a mixture comprising a duplex, a polymerase, and a cleaving reagent, the duplex comprising a growing nucleic acid strand and a nucleic acid strand to be sequenced, the duplex comprising at least one modified nucleotide of any one of claims 11-13; (b') performing a reaction comprising the following steps (i), (ii), and (iii), optionally, repeating one or more times: step (i): using the polymerase to incorporate the modified nucleotide into the growing nucleic acid strand to form a nucleic acid intermediate comprising a reversible blocking group and a detectable label: step (ii): detecting the nucleic acid intermediate; step (iii): using the cleaving reagent to cleave the reversible blocking group and the detectable label contained in the nucleic acid intermediate; Preferably, the cleaving reagent used for the cleavage of the reversible blocking group and the cleavage of the detectable label is the same reagent.

20. The method of claim 19, wherein, The method further has one or more of the technical features selected from the following (I)-(IX): (I) the duplex is immobilized on a support; (II) the growing nucleic acid strand is a primer; preferably, the primer forms the duplex by annealing to the nucleic acid strand to be sequenced; (III) the duplex, the modified nucleotide, and the polymerase together form a reaction system; (IV) using the polymerase to incorporate the modified nucleotide into the growing nucleic acid strand to form the nucleic acid intermediate comprising the reversible blocking group and the detectable label under conditions that allow the polymerase to perform nucleotide polymerization; and (V) the cleaving reagent is a chemical reagent; (V) removing the solution phase of the reaction mixture of the previous step before any one of the steps of detecting the nucleic acid intermediate; (VI) contacting the excision reagent with the duplex or the growing nucleic acid strand in the reaction mixture; (VII) the excision reagent is capable of excising the reversible blocking group and the detectable label contained in the modified nucleotide incorporated into the growing nucleic acid strand, and does not affect the phosphodiester bond on the duplex backbone; (VIII) removing the solution phase of the reaction mixture of this step after any one of the steps of excising the reversible blocking group and the detectable label contained in the nucleic acid intermediate; (IX) after step (ii), further comprising: determining the type of the modified nucleotide incorporated into the growing nucleic acid strand in step (i) based on the signal detected in step (ii), and determining the type of the nucleotide at the corresponding position in the nucleic acid strand to be sequenced based on the principle of base pairing complementarity.

21. Use of the compound of any one of claims 1-8 or a stereoisomer, tautomer, crystal form, salt, ester, or solvate thereof in sequencing, expression analysis, hybridization analysis, genetic analysis, RNA analysis, protein binding assay, in vitro diagnosis, immunoassay, molecular marker field. Preferably, the molecular marker is used for cell imaging, tissue imaging, or imaging of living organisms.

22. Use of the compound of any one of claims 1-8 or a stereoisomer, tautomer, crystal form, salt, ester, or solvate thereof in fluorescent labeling, quantification, or detection of proteins, enzymes, or nucleic acids.

23. Use of the nucleoside, nucleotide, or oligonucleotide of any one of claims 11-13 or the kit of claim 14 in sequencing.

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