Methods, kits, and reactive supports for 3' labeling of oligonucleotides
AssigneeAbbott Laboratories
InventorJames R. Fino
Life of this patent
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Abstract
In a first aspect, the invention involves a reactive support useful for automated synthesis of oligonucleotides. The reactive support comprises a label moiety (e.g. hapten) covalently bonded via a stable bond to a trifunctional spacer. The labeled trifunctional spacer complex is covalently bonded to a solid support via a cleavable bond. One arm of the trifunctional spacer attaches the solid phase; another arm attaches the label; while the third arm provides a hydroxyl group useful for synthesizing a labeled oligonucleotide. Upon synthesis, the cleavable bond is broken, yielding the labeled oligonucleotide. Methods for labeling oligonucleotides and useful kits are also described.
Claims27
A reactive support having a free or protected hydroxyl group useful for synthesis of an oligonucleotide, said support comprising a label moiety covalently bonded via a stable bond to a trifunctional spacer containing the hydroxyl group to form a labeled linker complex, the labeled linker complex being covalently bonded to a solid support via a cleavable bond.
A reactive support having a free or protected hydroxyl group useful for synthesis of an oligonucleotide, said support comprising a label moiety covalently bonded via a stable bond to a trifunctional spacer containing the hydroxyl group to form a labeled linker complex, the labeled linker complex being covalently bonded to a solid support via a cleavable bond;
- wherein said trifunctional spacer is a cyclic or heterocyclic molecule.
A process of preparing a reactive support having a label and a hydroxyl group useful for synthesizing a labeled oligonucleotide, said process comprising:
- a. reacting the second functionality of a trifunctional linker having three functionalities, a first one of whose functionalities is a hydroxyl group or a protected hydroxyl group and each of whose functionalities has or can be made to have a differential reactivity, with a label moiety having a reactive group capable of reacting with the second functionality to form a stable bond connecting the label moiety to the trifunctional linker;
- and
- b. reacting the third functionality of the trifunctional linker with one or more reagents capable of imparting a cleavable bond to the trifunctional linker, and with a functionalized solid support under conditions such that the trifunctional linker is covalently attached to the solid support via the cleavable bond.
A method for labeling the 3' end of an oligonucleotide synthesized on a solid support, said method comprising:
- a. reacting the second functionality of a trifunctional linker having three functionalities, a first one of whose functionalities is a protected hydroxyl group and each of whose functionalities has or can be made to have a differential reactivity, with a label moiety having a reactive group capable of reacting with the second functionality to form a stable bond connecting the label moiety to the trifunctional linker;
- b. reacting the third functionality of the trifunctional linker with one or more reagents capable of imparting a cleavable bond to the trifunctional linker, and with a functionalized solid support under conditions such that the trifunctional linker is covalently attached to the solid support via the cleavable bond.
- c. deprotecting the hydroxyl group;
- and
- d. synthesizing an oligonucleotide from the deprotected hydroxyl group.
Dependencies are inferred from the claim text.