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US 12,753,445 B2Granted
Composition and methods related to modification of 5 hydroxymethylcytosine (5-hmC)
AssigneeThe University of Chicago
InventorsChuan He, Chunxiao Song
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Abstract
The present invention relates generally to the field of molecular biology. More particularly, it concerns methods and compositions for detecting, evaluating, and/or mapping 5-hydroxymethyl-modified cytosine bases within a nucleic acid molecule.
Claims19
A method for labeling 5-hydroxymethylcytosines in a nucleic acid molecule, the method comprising:
- placing in a reaction mixture:
- the nucleic acid molecule;
- a β-glucosyltransferase;
- a synthetically modified uridine diphosphoglucose (UDP-Glu) molecule comprising a modified glucose molecule comprising either an azide functional group or a thiol functional group;
- and
- a label molecule comprising a moiety capable of attaching to the functional group on the modified glucose molecule;
- and
- labeling 5-hydroxymethylcytosines in the nucleic acid molecule in the reaction mixture.
5 more elements
A method for labeling 5-hydroxymethylcytosines in a nucleic acid molecule, the method comprising:
- placing in a reaction mixture:
- the nucleic acid molecule;
- a β-glucosyltransferase;
- and
- a synthetically modified uridine diphosphoglucose (UDP-Glu) molecule comprising a modified glucose molecule comprising, on carbon-6, an azide moiety,
- under conditions sufficient to glycosylate 5-hydroxymethylcytosines in the nucleic acid molecule with the modified glucose molecule;
- and
- placing in the reaction mixture a dibenzocyclooctyne-PEG4-biotin conjugate under conditions sufficient to attach the dibenzocyclooctyne-PEG4-biotin conjugate to the azide moiety on the modified glucose via Click chemistry, thereby labeling 5-hydroxymethylcytosines in a nucleic acid.
5 more elements
Dependencies are inferred from the claim text.