Propargylethoxyamino nucleotides
View Patent ↗Propargylethoxyamino nucleosides are disclosed having the structure wherein R 1 and R 2 are —H, lower alkyl, or label; B is a 7-deazapurine, purine, or pyrimidine nucleoside base; W 1 is —H or —OH; W 2 is —OH or a moiety which renders the nucleoside incapable of forming a phosphodiester bond at the 3′-position; and W 3 is —PO 4 , —P 2 O 7 , —P 3 O 10 , phosphate analog, or —OH. Additionaly, a primer extension method is provided employing the above propargylethoxyamino nucleosides.
1. A method for performing a primer extension reaction comprising:
annealing a polynucleotide primer to a portion of a template nucleic acid, and
extending the annealed primer in the presence of a compound having the structure:
wherein:
n is 1 or 2,
R1 and R2 taken separately are each selected from —H, lower alkyl, protecting group and label,
B is a 7-deazapurine, purine, or pyrimidine nucleoside base,
wherein when B is purine or 7-deazapurine, the sugar moiety is attached at the N9-position of the purine or deazapurine, and when B is pyrimidine, the sugar moiety is attached at the N1-position of the pyrimidine, and
wherein if B is a purine, the adjacent triple-bonded carbon is attached to the 8-position of the purine, if B is 7-deazapurine, the adjacent triple-bonded carbon is attached to the 7-position of the 7-deazapurine, and if B is pyrimidine, the adjacent triple-bonded carbon is attached to the 5-position of the pyrimidine,
W1 is selected from —H and —OH,
W2 is —OH or a moiety which renders the nucleoside incapable of forming a phosphodiester bond at the 3′-position, and
W3 is selected from —PO4, —P2O7, —P3O10, phosphate analog, and —OH,
whereby an extended primer is formed comprising at least one of the compound.
2. The method of claim 1 wherein one of R1 and R2 is label and the other is —H.
3. The method of claim 2 wherein W 3 is —P 3 O 10 .
4. The method of claim 3 wherein W 1 is H.
5. The method of claim 4 wherein W 2 is —OH or a moiety selected from —H, azido, amino, fluoro and methoxy.
6. The method of claim 3 wherein W 2 is —OH or a moiety selected from —H, azido, amino, fluoro and methoxy.
7. The method of claim 2 wherein W 1 is H.
8. The method of claim 7 wherein W 2 is —OH or a moiety selected from —H, azido, amino, fluoro and methoxy.
9. The method of claim 2 wherein W 2 is —OH or a moiety selected from —H, azido, amino, fluoro and methoxy.
10. The method of claim 1 wherein W 3 is —P 3 O 10 .
11. The method of claim 10 wherein W 1 is H.
12. The method of claim 11 wherein W 2 is —OH or a moiety selected from —H, azido, amino, fluoro and methoxy.
13. The method of claim 10 wherein W 2 is —OH or a moiety selected from —H, azido, amino, fluoro and methoxy.
14. The method of claim 1 wherein W 1 is H.
15. The method of claim 14 wherein W 2 is —OH or a moiety selected from —H, azido, amino, fluoro and methoxy.
16. The method of claim 1 wherein W 2 is —OH or a moiety selected from —H, azido, amino, fluoro and methoxy.
17. The method of claim 1 wherein R 1 is a label selected from a rhodamine dye and a fluorescein dye.
18. The method of claim 17 wherein W 1 is —H and W 2 is —H.
19. The method of claim 18 wherein the label is a rhodamine dye.
20. The method of claim 1 wherein the extending is mediated by a DNA polymerase.