PROPARGYL SUBSTITUTED NUCLEOSIDE COMPOUNDS AND METHODS
First Claim
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1. A nucleotide comprising a nucleobase disposed between a sugar or sugar analog and a propargylic moiety, wherein the nucleobase comprises a pyrimidine nucleobase or pyrimidine nucleobase analog, the propargylic moiety is covalently attached to the nucleobase at the C-5 position of the nucleobase, the sugar or sugar analog is covalently attached to the nucleobase at the N−
- 1 position of the nucleobase, the sugar or sugar analog comprises a 5′
-monophosphate group, 5′
-diphosphate group, or 5′
-triphosphate group, wherein the propargylic moiety comprises(i) a propargyl group covalently attached by its methylene carbon to the nucleobase,(ii) a detectable label, and(iii) a linker covalently linking the acetylene carbon of the propargyl group to the detectable label.
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Abstract
The present teachings relate to nucleobase, nucleoside and nucleotide compounds, methods of synthesis, and uses thereof. The present teachings provide compounds, such as nucleobase, nucleoside and/or nucleotide compounds including a propargyl linker, and methods for making or using such compounds.
9 Citations
108 Claims
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1. A nucleotide comprising a nucleobase disposed between a sugar or sugar analog and a propargylic moiety, wherein the nucleobase comprises a pyrimidine nucleobase or pyrimidine nucleobase analog, the propargylic moiety is covalently attached to the nucleobase at the C-5 position of the nucleobase, the sugar or sugar analog is covalently attached to the nucleobase at the N−
- 1 position of the nucleobase, the sugar or sugar analog comprises a 5′
-monophosphate group, 5′
-diphosphate group, or 5′
-triphosphate group, wherein the propargylic moiety comprises(i) a propargyl group covalently attached by its methylene carbon to the nucleobase, (ii) a detectable label, and (iii) a linker covalently linking the acetylene carbon of the propargyl group to the detectable label. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 102)
- 1 position of the nucleobase, the sugar or sugar analog comprises a 5′
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24. A labeled nucleoside comprising a nucleobase disposed between a sugar or a sugar analog and a propargylic moiety, wherein the nucleobase is selected from a pyrimidine base or a pyrimidine base analog, the propargylic moiety is covalently attached to the nucleobase at the C-5 position of the nucleobase, the sugar or sugar analog is covalently attached to the nucleobase at the N−
- 1 position of the nucleobase, and wherein the propargylic moiety comprises
(i) a propargyl group covalently attached by its methylene carbon to the nucleobase, (ii) a detectable label, and (iii) a linker covalently linking the acetylene carbon of the propargyl group to the detectable label. - View Dependent Claims (25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46)
- 1 position of the nucleobase, and wherein the propargylic moiety comprises
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47. A polynucleotide comprising at least one nucleobase comprising the structure
NUC—- CH2—
≡
—
X-LABELwherein NUC comprises a pyrimidine nucleobase or pyrimidine nucleobase analog, X comprises a bond or a linker having from 1-50 chain atoms selected from C, N, O, S, P and Si, and LABEL comprises at least one detectable label. - View Dependent Claims (48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69)
- CH2—
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53. The polynucleotide of claim 47 wherein the linker comprises from 1-50 atoms selected from C, H, N, O, S, P and Si.
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54. The polynucleotide of claim 47 wherein the linker comprises at least one anionic moiety.
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55. The polynucleotide of claim 47 wherein the linker comprises at least one cationic moiety.
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56. The polynucleotide of claim 47 wherein the linker comprises at least one amide linkage.
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57. The polynucleotide of claim 56 wherein the linker comprises at least one diradical selected from
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58. The polynucleotide of claim 56 wherein the linker comprises at least one diradical selected from
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59. The polynucleotide of claim 58 wherein Z is unsubstituted amine.
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60. The polynucleotide of claim 58 wherein Z is substituted amine.
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61. The polynucleotide of claim 47 wherein the detectable label comprises at least one of a fluorescent dye, an energy transfer dye, a quencher or biotin.
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62. The polynucleotide of claim 47 wherein the detectable label comprises a fluorescent dye selected from a rhodamine dye, a fluorescein dye, a rhodol dye, an energy transfer dye, a cyanine dye, a phthalocyanine and a squaraine.
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63. The polynucleotide of claim 61 wherein the fluorescent dye comprises a fluorescein dye, a rhodamine dye or an energy transfer dye.
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64. The polynucleotide of claim 47 wherein the detectable label comprises an energy transfer dye comprising a donor dye covalently attached to an acceptor dye, wherein either the donor dye or the acceptor dye is covalently attached to the linker, and the acceptor dye is capable of absorbing the light emitted by the donor dye.
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65. The polynucleotide of claim 64 wherein the donor dye is covalently attached to the linker.
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66. The polynucleotide of claim 64 wherein the donor dye comprises a fluorescein dye or a rhodamine dye.
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67. The polynucleotide of claim 66 wherein the acceptor dye comprises one of a rhodamine dye, a fluorescein dye, a cyanine dye or a non-fluorescent quencher.
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68. The polynucleotide of claim 67 wherein the donor dye is covalently linked to the acceptor dye through a linker-moiety that is different from the first said linker.
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69. The polynucleotide of claim 64 wherein the donor dye comprises a fluorescein or a rhodamine dye and the acceptor dye is a non-fluorescent quencher.
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70. A nucleoside comprising a nucleobase disposed between a sugar or a sugar analog and a propargylic moiety, wherein the nucleobase is selected from a pyrimidine base or a pyrimidine base analog, the propargylic moiety is covalently attached to the nucleobase at the C-5 position of the nucleobase, the sugar or sugar analog is covalently attached to the nucleobase at the N−
- 1 position of the nucleobase, and wherein the propargylic moiety comprises a propargyl group covalently attached to the nucleobase at the methylene carbon of the propargyl group.
- View Dependent Claims (71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83)
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84. A nucleotide comprising a nucleobase disposed between a sugar or a sugar analog and a propargylic moiety, wherein the nucleobase is selected from a pyrimidine base or a pyrimidine base analog, the propargylic moiety is covalently attached to the nucleobase at the C-5 position of the nucleobase, the sugar or sugar analog is covalently attached to the nucleobase at the N−
- 1 position of the nucleobase, the sugar or sugar analog comprises a 5′
-monophosphate group, 5′
-diphosphate group or 5′
-triphosphate group and wherein the propargylic moiety comprises a propargyl group covalently attached to the nucleobase at the methylene carbon of the propargyl group. - View Dependent Claims (85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96)
- 1 position of the nucleobase, the sugar or sugar analog comprises a 5′
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97. A method of extending a polynucleotide comprising
forming a hybridization complex between a 3′ - -extendable polynucleotide and a complementary sequence in a polynucleotide, and
extending the 3′
-extendable polynucleotide in the presence of a nucleotide 5′
-triphosphate and a template-dependent nucleic acid polymerase under conditions effective to append the nucleotide 5′
triphosphate to the 3′
-end of the extendable polynucleotide, wherein the nucleotide 5′
-triphosphate comprises nucleobase disposed between a sugar or sugar analog and a propargylic moiety, the nucleobase comprises a pyrimidine nucleobase or pyrimidine nucleobase analog, the propargylic moiety is covalently attached to the nucleobase at the C-5 position of the nucleobase, the sugar or sugar analog is covalently attached to the nucleobase at the N−
1 position of the nucleobase, the sugar or sugar analog comprises a 5′
-monophosphate group, a 5′
-diphosphate group, or a 5′
-triphosphate group, and the propargylic moiety comprises (i) a propargyl group covalently attached by its methylene carbon to the nucleobase, (ii) a detectable label, and (iii) a linker covalently linking the acetylene carbon of the propargyl group to the detectable label. - View Dependent Claims (98, 99, 100, 101)
- -extendable polynucleotide and a complementary sequence in a polynucleotide, and
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103. A method for making a thymidine analog comprising:
contacting a protected or unprotected 5-halomethyl uridine nucleotide with a copper acetylide to form an adduct comprising a 5-propargyl uridine. - View Dependent Claims (104)
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105. A compound having the formula:
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NUC—
CH2—
C≡
C—
X-LABELwherein NUC comprises a nucleoside comprising a sugar moiety and a nucleobase selected from a pyrimidine nucleobase or a pyrimidine nucleobase analog, the CH2 moiety is covalently attached to the 5-atom of the nucleobase, X is bond or a linker having from 1 to 50 chain atoms each selected from C, H, N, O, S, P and Si, and the sugar moiety is covalently attached to the N−
1 position of the nucleobase.- View Dependent Claims (106, 107)
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108. A compound comprising the structure
Specification