Fluorescent dye compounds, conjugates and uses thereof

• US 7,256,292 B2
• Filed: 09/16/2005
• Issued: 08/14/2007
• Est. Priority Date: 09/16/2004
• Status: Active Grant

• Current Assignee: Applied Biosystems LLC (Thermo Fisher Scientific Incorporated)
• Original Assignee: Applera Corporation (Thermo Fisher Scientific Incorporated)
• Inventors: Benson, Scott C., Graham, Ronald J., Zou, Ruiming, Upadhya, Krishna G.
• Primary Examiner(s): AULAKH, CHARANJIT
• Application Number:
  US11/229,012
• Publication Number:
  US 20060167257A1
• Time in Patent Office: 697 Days
• Field of Search: 546/102, 546/61, 546/58
• US Class Current: 546/102
• CPC Class Codes: C07D 219/06 Oxygen atoms
• 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: C09B 15/00 Acridine dyes
• CPC Class Codes: C09K 11/06 containing organic luminescent materials
• CPC Class Codes: C09K 2211/1011 Condensed systems
• CPC Class Codes: C09K 2211/1029 containing one nitrogen atom as the heteroatom
• CPC Class Codes: C12Q 1/6816 characterised by the detection means C12Q1/6804 takes precedence
• CPC Class Codes: C12Q 1/6869 Methods for sequencing
• CPC Class Codes: G01N 33/582 with fluorescent label
• CPC Class Codes: Y10S 436/80 Fluorescent dyes, e.g. rhodamine
• CPC Class Codes: Y10T 436/143333 Saccharide [e.g., DNA, etc.]

Abstract

The present teachings generally relate to fluorescent dyes, linkable forms of fluorescent dyes, energy transfer dyes, reagents labeled with fluorescent dyes and uses thereof.

Claims

1. A compound having a structure selected from

2. The compound of claim 1, wherein R₆ and R₇ are each independently selected from fluorine, chlorine and bromine.

3. The compound of claim 2, wherein R₆ and R₇ are fluorine.

4. The compound of claim 2, wherein R₆ and R₇ are chlorine.

5. The compound of claim 2, wherein R₆ and R₇ are bromine.

6. The compound of claim 1, wherein R₆ is —

H and R₇ is selected from —

H, —

CO₂H, —

CO₂R, —

SO₃H, —

SO₃R, —

CH₂CO₂H, —

CH₂CO₂R, —

CH₂SO₃H, —

CH₂SO₃R, —

CH₂NH₂, and —

CH₂NHR.

7. The compound of claim 1, wherein R₆ is —

H and R₇ is selected from —

H, —

SO₃H, —

SO₃R, —

CH₂CO₂H, —

CH₂CO₂R, —

CH₂NH₂, and —

CH₂NHR.

8. The compound of claim 1, having structure (I), wherein R₆ and R₇ are each independently selected from fluorine, chlorine and bromine and R₂ is selected from —

CO₂H, —

CO₂R, —

SO₃H, —

SO₃R, —

CH₂CO₂H, —

CH₂CO₂R, —

CH₂SO₃H, —

CH₂SO₃R, —

CH₂NH₂, —

CH₂NHR and —

NO₂.

9. The compound of any one of claims 1-8, wherein the compound has the structure (I) and R₂ is selected from —

SO₃H, —

SO₃R, —

CH₂NH₂, —

CH₂NHR, and —

NO₂.

10. The compound of any one of claims 1-8, wherein the compound has the structure (I) and R₂ is selected from —

SO₃H and —

SO₃R.

11. The compound of any one of claims 1-8, wherein the compound has the structure (I) and R₂ is selected from —

CH₂NH₂ and —

CH₂NHR.

12. The compound of claim 9, wherein R₂ is —

SO₃H.

13. The compound of claim 9, wherein R₂ is —

SO₃R.

14. The compound of claim 9, wherein R₂ is —

CH₂NH₂.

15. The compound of claim 9, wherein R₂ is —

CH₂NHR.

16. The compound of any one of claims 1-8, wherein R is

17. The compound of any one of claims 1, 6 or 7, wherein the compound has the structure (I), R₇ is —

SO₃H and R₂ is —

CH₂NH₂.

18. The compound of any one of claims 1, 6 or 7, wherein the compound has the structure (I), R₇ is —

SO₃H and R₂ is —

CH₂NHR.

19. The compound of claim 18, wherein R is substituted benzoyl.

20. The compound of claim 19, wherein R is

21. The compound of any one of claims 1-8, wherein R₄ and R₅ are methyl.

22. The compound of any one of claims 1-8, wherein R₁ and R₃ are —

H.

23. The compound of claim 1, having structure (II), wherein R₉ is selected from —

H, —

CO₂H, —

CO₂R, —

SO₃H, —

SO₃R, —

CH₂CO₂H, —

CH₂CO₂R, —

CH₂SO₃H, —

CH₂SO₃R, —

CH₂NH₂, —

CH₂NHR and —

NO₂, and R₃ and R₈-R₁₀ are —

H.

24. The compound of claim 23, wherein R₉ is selected from —

SO₃H, —

SO₃R, —

CH₂NH₂, —

CH₂NHR, and —

NO₂.

25. The compound of claim 23, wherein R₉ is selected from —

SO₃H and —

SO₃R.

26. The compound of claim 23, wherein R₉ is selected from —

CH₂NH₂ and —

CH₂NHR.

27. The compound of claim 23, wherein R₉ is —

SO₃H.

28. The compound of claim 23, wherein R₉ is —

SO₃R.

29. The compound of claim 23, wherein R₉ is —

CH₂NH₂.

30. The compound of claim 23, wherein R₉ is —

CH₂NHR.

31. The compound of any one of claims 23-26, 28 or 30, wherein R is

32. The compound of claim 6, having structure (II), wherein R₉ is selected from —

H, —

CO₂H, —

CO₂R, —

SO₃H, —

SO₃R, —

CH₂CO₂H, —

CH₂CO₂R, —

CH₂SO₃H, —

CH₂SO₃R, —

CH₂NH₂, —

CH₂NHR and —

NO₂, and R₃ and R₁₀-R₁₂ are —

H.

33. The compound of claim 32, wherein R₇ is —

SO₃H and R₉ is —

CH₂NH₂.

34. The compound of claim 32, wherein R₇ is —

SO₃H and R₉ is —

CH₂NHR.

35. The compound of claim 32, wherein R is

36. The compound of any claim 1, having structure (II), wherein R₁₀ is selected from —

H, —

CO₂H, —

CO₂R, —

SO₃H, —

SO₃R, —

CH₂CO₂H, —

CH₂CO₂R, —

CH₂SO₃H, —

CH₂SO₃R, —

CH₂NH₂, —

CH₂NHR and —

NO₂, and R₃, R₉ and R₁₁-R₁₂ are —

H.

37. The compound of claim 36, wherein R₁₀ is selected from —

SO₃H, —

SO₃R, —

CH₂NH₂, —

CH₂NHR, and —

NO₂.

38. The compound of claim 36, wherein R₁₀ is selected from —

SO₃H and —

SO₃R.

39. The compound of claim 36, wherein R₁₀ is selected from —

CH₂NH₂ and —

CH₂NHR.

40. The compound of claim 36, wherein R₁₀ is —

SO₃H.

41. The compound of claim 36, wherein R₁₀ is —

SO₃R.

42. The compound of claim 36, wherein R₁₀ is —

CH₂NH₂.

43. The compound of claim 36, wherein R₁₀ is —

CH₂NHR.

44. The compound of any one of claims 36-39, 41 or 43, wherein R is

45. The compound of claim 6, having structure (II), wherein R₁₀ is selected from —

H, —

CO₂H, —

CO₂R, —

SO₃H, —

SO₃R, —

CH₂CO₂H, —

CH₂CO₂R, —

CH₂SO₃H, —

CH₂SO₃R, —

CH₂NH₂, —

CH₂NHR and —

NO₂ and R₃, R₉ and R₁₁-R₁₂ are —

H.

46. The compound of claim 45, wherein R₇ is —

SO₃H and R₁₀ is —

CH₂NH₂.

47. The compound of claim 45, wherein R₇ is —

SO₃H and R₁₀ is —

CH₂NHR.

48. The compound of claim 47, wherein R is

49. The compound of claim 1, having structure (II), wherein R₁₁ is selected from —

H, —

CO₂H, —

CO₂R, —

SO₃H, —

SO₃R, —

CH₂CO₂H, —

CH₂CO₂R, —

CH₂SO₃H, —

CH₂SO₃R, —

CH₂NH₂, —

CH₂NHR and —

NO₂, and R₃, R₉-R₁₀ and R₁₂ are —

H.

50. The compound of claim 49, wherein R₁₁ is selected from —

SO₃H, —

SO₃R, —

CH₂NH₂, —

CH₂NHR, and —

NO₂.

51. The compound of claim 49, wherein R₁₁ is selected from —

SO₃H and —

SO₃R.

52. The compound of claim 49, wherein R₁₁ is selected from —

CH₂NH₂ and —

CH₂NHR.

53. The compound of claim 49, wherein R₁₁ is —

SO₃H.

54. The compound of claim 49, wherein R₁₁ is —

SO₃R.

55. The compound of claim 49, wherein R₁₁ is —

CH₂NH₂.

56. The compound of claim 49, wherein R₁₁ is —

CH₂NHR.

57. The compound of any one of claims 49-52, 54 or 56, wherein R is

58. The compound of claim 6, having structure (II), wherein R₁₁ is selected from —

H, —

CO₂H, —

CO₂R, —

SO₃H, —

SO₃R, —

CH₂CO₂H, —

CH₂CO₂R, —

CH₂SO₃H, —

CH₂SO₃R, —

CH₂NH₂, —

CH₂NHR and —

NO₂, and R₃, R₉-R₁₀ and R₁₂ are —

H.

59. The compound of claim 58, wherein R₇ is —

SO₃H and R₁₁ is —

CH₂NH₂.

60. The compound of claim 58, wherein R₇ is —

SO₃H and R₁₁ is —

CH₂NHR.

61. The compound of claim 60, wherein R is

62. The compound of claim 1, having structure (II), wherein R₁₂ is selected from —

H, —

CO₂H, —

CO₂R, —

SO₃H, —

SO₃R, —

CH₂CO₂H, —

CH₂CO₂R, —

CH₂SO₃H, —

CH₂SO₃R, —

CH₂NH₂, —

CH₂NHR and —

NO₂, and R₃ and R₉-R₁₁ are —

H.

63. The compound of claim 62, wherein R₁₂ is selected from —

SO₃H, —

SO₃R, —

CH₂NH₂, —

CH₂NHR, and —

NO₂.

64. The compound of claim 62, wherein R₁₂ is selected from —

SO₃H and —

SO₃R.

65. The compound of claim 62, wherein R₁₂ is selected from —

CH₂NH₂ and —

CH₂NHR.

66. The compound of claim 62, wherein R₁₂ is —

SO₃H.

67. The compound of claim 62, wherein R₁₂ is —

SO₃R.

68. The compound of claim 62, wherein R₁₂ is —

CH₂NH₂.

69. The compound of claim 62, wherein R₁₂ is —

CH₂NHR.

70. The compound of any one of claims 62-65, 67 or 69, wherein R is

71. The compound of claim 6, having structure (II), wherein R₁₂ is selected from —

H, —

CO₂H, —

CO₂R, —

SO₃H, —

SO₃R, —

CH₂CO₂H, —

CH₂CO₂R, —

CH₂SO₃H, —

CH₂SO₃R, —

CH₂NH₂, —

CH₂NHR and —

NO₂, and R₃ and R₉-R₁₁ are —

H.

72. The compound of claim 71, wherein R₇ is —

SO₃H and R₁₂ is —

CH₂NH₂.

73. The compound of claim 71, wherein R₇ is —

SO₃H and R₁₂ is —

CH₂NHR.

74. The compound of claim 73, wherein R is

75. The compound of any one of claims 23, 32, 36, 45, 49, 58, 62 or 71, wherein R₉-R₁₂ are H.

76. The compound of any one of claims 23, 32, 36, 45, 49, 58, 62 or 71, wherein R₄ and R₅ are methyl.

77. The compound according to any one of claims 1-8, wherein R₂ and R₃ are —

NO₂ and R₄ and R₅ are methyl.

78. The compound according to any one of claims 1, 6 or 7, having structure (III) and R₁ and R₃ are —

H.

79. The compound of claim 78, wherein R₆ is —

H.

80. The compound according to claim 78, wherein R₁₃, R₁₅ and R₁₆ are —

H.

81. The compound of claim 80, wherein R₂ is —

SO₃H.

82. The compound of claim 80, wherein R₂ is —

H.

83. The compound according to claim 80, wherein R₁₄ is —

CH₂NH₂.

84. The compound according to any one of claims 80-82, wherein R₁₄ is —

CH₂NHR.

85. The compound of claim 84, wherein R is

86. The compound of claim 81, wherein R₁₄ is —

H.

87. The compound of claim 80, wherein R₁₄ is —

SO₃H.

88. The compound of claim 80, wherein R₁₄ is —

H.

89. The compound according to claim 80, wherein R₂ is —

CH₂NH₂.

90. The compound according to claim 80, wherein R₂ is —

CH₂NHR.

91. The compound of claim 90, wherein R is

92. The compound of claim 87, wherein R₂ is —

H.

93. The compound according to claim 89, wherein R₄ and R₅ are methyl.

94. The compound according to any one of claims 1, 6 or 7, wherein the compound has structure (IV).

95. The compound of claim 94, wherein R₁₄ is —

SO₃H.

96. The compound of claim 94, wherein R₁₄ is —

H.

97. The compound according to claim 94, wherein R₁₀ is —

CH₂NH₂.

98. The compound according to claim 94, wherein R₁₀ is —

CH₂NHR.

99. The compound of claim 98, wherein R is

100. The compound according to claim 94, wherein R₁₄ is —

H.

101. The compound according to claim 94, wherein R₃, R₆, R₉, R₁₁, R₁₂, R₁₃, R₁₅ and R₁₆ are —

H.

102. The compound according to claim 94, wherein R₄ and R₅ are methyl.

103. A compound of claim 1 having the structure:

104. A compound of claim 1 having the structure:

105. A compound of claim 1 having the structure:

106. A compound of claim 1 having the structure

107. A compound of claim 1 having the structure:

108. A compound of claim 1 having the structure:

109. A compound of claim 1 having the structure:

110. A compound of claim 1 having the structure:

111. A compound of claim 1 having the structure:

112. A compound of claim 1 having the structure:

113. A compound of claim 1 having the structure:

114. The compound of claim 82, wherein R14 is H.