Nucleotide analogues
The present invention provides methods, compositions, mixtures and kits utilizing deoxynucleoside triphosphates comprising a 3′-O position capped by a group comprising methylenedisulfide as a cleavable protecting group and a detectable label reversibly connected to the nucleobase of said deoxynucleoside. Such compounds provide new possibilities for future sequencing technologies, including but not limited to Sequencing by Synthesis.
1. A labeled deoxynucleoside triphosphate according to the following structure:
wherein D is selected from the group consisting of an azide, disulfide alkyl, disulfide substituted alkyl groups; wherein said label is a dye and B is a nucleobase.
2. A labeled deoxynucleoside triphosphate according to the following structure:
wherein D is selected from the group consisting of an azide, disulfide alkyl, disulfide substituted alkyl groups; wherein said label is a dye and B is a nucleobase.
3. A labeled deoxynucleoside triphosphate according to the following structure:
wherein D is selected from the group consisting of an azide, disulfide alkyl, disulfide substituted alkyl groups; wherein said label is a dye and B is a nucleobase.
4. The labeled deoxynucleoside triphosphate according to claim 2 , wherein the compound has the structure:
5. The labeled deoxynucleoside triphosphate according to claim 1 , wherein the compound has the structure:
6. The labeled deoxynucleoside triphosphate according to claim 3 , wherein the compound has the structure:
7. The labeled deoxynucleoside triphosphate according to claim 1 , wherein the compound has the structure:
8. The labeled deoxynucleoside triphosphate according to claim 1 , wherein the compound has the structure:
9. The labeled deoxynucleoside triphosphate according to claim 1 , wherein the compound has the structure:
10. The labeled deoxynucleoside triphosphate according to claim 1 , wherein the compound has the structure:
11. The labeled deoxynucleoside triphosphate according to claim 1 , wherein the compound has the structure:
12. The labeled deoxynucleoside triphosphate according to claim 1 , wherein the compound has the structure:
13. The labeled deoxynucleoside triphosphate according to claim 1 , wherein the compound has the structure:
14. The labeled deoxynucleoside triphosphate according to claim 1 , wherein the compound has the structure:
15. The labeled deoxynucleoside triphosphate according to claim 2 , wherein the compound has the structure:
16. The labeled deoxynucleoside triphosphate according to claim 3 , wherein the compound has the structure:
17. The labeled deoxynucleoside triphosphate according to claim 2 , wherein the compound has the structure:
18. The labeled deoxynucleoside triphosphate according to claim 1 , wherein the compound has the structure:
19. The labeled deoxynucleoside triphosphate according to claim 3 , wherein the compound has the structure:
20. A kit comprising a DNA polymerase and at least one deoxynucleoside triphosphate according to the following structure:
wherein D is selected from the group consisting of an azide, disulfide alkyl, disulfide substituted alkyl groups, disulfide allyl, and disulfide substituted allyl groups; B is a nucleobase; A is an attachment group selected from the group consisting of exocyclic amine, propargyl amine, and propargyl hydroxyl; C is a cleavable site core selected from the group consisting of:
and,
wherein R 1 and R 2 are independently selected alkyl groups; L 1 and L 2 , are connecting groups; and Label is a label selected from the group consisting of fluorophore dyes, energy transfer dyes, mass-tags, biotin, and haptenes.
21. A reaction mixture comprising a nucleic acid template with a primer hybridized to said template, a DNA polymerase and at least one deoxynucleoside triphosphate according to the following structure:
wherein D is selected from the group consisting of an azide, disulfide alkyl, disulfide substituted alkyl groups, disulfide allyl, and disulfide substituted allyl groups; B is a nucleobase; A is an attachment group selected from the group consisting of exocyclic amine, propargyl amine, and propargyl hydroxyl; C is a cleavable site core selected from the group consisting of:
wherein R 1 and R 2 are independently selected alkyl groups; L 1 and L 2 , are connecting groups; and Label is a label selected from the group consisting of fluorophore dyes, energy transfer dyes, mass-tags, biotin, and haptenes.
22. The reaction mixture of claim 21 , wherein the template, primer, polymerase a deoxynucleoside triphosphate are in a solution.
23. The reaction mixture of claim 22 , wherein said solution is a buffer.