Method for deblocking of labeled oligonucleotides
View Patent ↗The invention relates to a process for deblocking substantially a blocked, detectably labeled oligonucleotide by contacting the blocked detectably labeled oligonucleotide with an effective amount of a nucleophilic amino compound under conditions that result in substantial deblocking of the oligonucleotide, thereby giving the substantially deblocked oligonucleotide.
1. A composition in a sealable chamber comprising a blocked, labeled oligonucleotide, an effective amount of a nucleophilic amino compound sufficient to substantially deblock said oligonucleotide and one or more carbohydrates and/or sugars, said composition being at a pressure greater than atmospheric pressure.
2. A process for substantially deblocking a blocked, labeled oligonucleotide comprising contacting, in a sealable chamber, the blocked labeled oligonucleotide at a pressure greater than atmospheric pressure with an effective amount of a nucleophilic amino compound in the presence of one or more carbohydrates and/or sugars, thereby giving a substantially deblocked oligonucleotide.
3. The process of claim 2 , wherein the one or more carbohydrates and/or sugars are selected from the group consisting of sucrose and trehalose.
4. The process of claim 2 , wherein the label is a fluorescent label.
5. The process of claim 2 , wherein the fluorescent label is hexachlorofluorescein.
6. The process of claim 2 , wherein the fluorescent label is 4-([4′-(dimethylamino)phenyl]azo)benzoic acid.
7. The process of claim 2 , wherein the nucleophilic amino compound is ammonia.
8. The process of claim 2 , wherein the nucleophilic amino compound is ammonia vapors.
9. The process of claim 2 , wherein the nucleophilic amino compound is a C 1-6 alkylamine.
10. The process of claim 2 , further comprising dissolving the substantially deblocked oligonucleotide in a buffer.
11. The process of claim 2 , wherein the conditions comprise carrying out the process at about room temperature to about 150° C.
12. The process of claim 2 , wherein the conditions comprise carrying out the process at about 95° C.
13. The process of claim 2 , wherein the conditions comprise carrying out the process for about 1 minute to about 2 hours.
14. The process of claim 2 , wherein the conditions comprise carrying out the process for about 45 minutes.
15. The process of claim 2 , wherein the substantially blocked, labeled oligonucleotide is immobilized on a solid phase.
16. The process of claim 15 , wherein the substantially deblocked oligonucleotide is released from the solid phase under the conditions.
17. The process of claim 16 , wherein the substantially deblocked oligonucleotide is recovered by washing the solid phase with water or a buffer.
18. A process for substantially deblocking a blocked, labeled oligonucleotide comprising contacting, in a sealable chamber, the blocked labeled oligonucleotide with an effective amount of ammonia saturated with water vapor in the presence of one or more carbohydrates and/or sugars at about 80 pounds per square inch, 95° C., for about 45 minutes, thereby giving the substantially deblocked oligonucleotide.
19. The process of claim 18 , wherein the blocked, labeled oligonucleotide is immobilized on a solid phase.