Pro-thiol aryl azide labelling of nucleic acids
AssigneeVector Laboratories, Inc.
InventorsMark E. Westling, Steven G. Daniel
Life of this patent
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Abstract
Reagents and methods for multi-step labeling of nucleic acids allow the addition of relatively insoluble or unstable labels to nucleic acid in the final step. Nucleic acids can be stored as a stable intermediate capable of reacting with a label conjugated to a thiol-reactive group.
Claims15
A pro-thiol aryl azide for coupling a thiol group to a nucleic acid, said pro-thiol aryl azide having the structure:
- ##STR40## wherein, R.sub.1 to R.sub.4, which may be the same or different, are selected from the group consisting of H, alkyl, nitro, hydroxy, halide, carboxylic acid, methoxy, cyano, amino groups and salts thereof;
- wherein E comprises an amine and salts thereof, and a chain of C.sub.1 -C.sub.20 having chain constituents selected from the group consisting of alkyl, alkenyl, alkynyl heteroatoms and salts thereof, said chain constituents having substituents selected from the group consisting of H, C.sub.1 -C.sub.6 alkyl, aryl, ester, ether, amine, amide, halo groups and salts thereof;
- and wherein Y is a group capable of protecting the sulfur moiety from chemical reaction and removable so as to generate a thiol moiety.
A covalent conjugate of a nucleic acid and a thiol-reactive label having a thiol-reactive group given by the structure:
- ##STR44## wherein, R.sub.1 to R.sub.4, which may be the same or different, are selected from the group consisting of H, alkyl, nitro, hydroxy, halide, carboxylic acid, methoxy, cyano, amino groups and salts thereof;
- wherein E comprises an amine and salts thereof, and a chain of C.sub.1 -C.sub.20 having chain constituents selected from the group consisting of alkyl, alkenyl, alkynyl heteroatoms and salts thereof, said chain constituents having substituents selected from the group consisting of H, C.sub.1 -C.sub.6 alkyl, aryl, ester, ether, amine, amide, halo groups and salts thereof;
- wherein R.sub.9 is the thiolated conjugate of the thiol-reactive group in the thio-reactive label;
- and
- wherein L is selected from the group consisting of a ligand or a detectable label.
A pro-thiol aryl azide for coupling a thiol group to a nucleic acid, said pro-thiol aryl azide having the structure:
- ##STR46## wherein, R.sub.1 to R.sub.4, which may be the same or different, are selected from the group consisting of H, alkyl, nitro, hydroxy, halide, carboxylic acid, methoxy, cyano, amino groups and salts thereof;
- wherein E is:
- ##STR47## and Y is:
- ##STR48##
A covalent conjugate of a nucleic acid and a thiol-reactive label having a thiol-reactive group given by the structure:
- ##STR49## wherein, R.sub.1 to R.sub.4, which may be the same or different, are selected from the group consisting of H, alkyl, nitro, hydroxy, halide, carboxylic acid, methoxy, cyano, amino groups and salts thereof;
- wherein E is:
- ##STR50## wherein R.sub.9 is the thiolated conjugate of the thiol-reactive group in the thio-reactive label;
- and
- wherein L is selected from the group consisting of a ligand or a detectable label.
Dependencies are inferred from the claim text.