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US 2026/0168988 A1Published application
Genetically encoded reporter expression systems and methods of making and using same
AssigneeDuke University
InventorKathryn Meyer
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Abstract
Provided herein are compositions and methods for detection of N6-methyladenosine (m6A) methylation in live cells. The provided compositions include expression systems for detection of m6A methylation and identification of agents that modulate m6A methylation.
Claims41
An expression system comprising:
- (a) a first DNA construct comprising a nucleic acid sequence encoding a fusion protein, wherein the fusion protein comprises an N6-methyladenosine (m6A) binding domain of a YT521-B homology (YTH) domain-containing protein fused to a catalytic domain of a cytidine deaminase or a catalytic domain of an adenosine deaminase;
- and
- (b) a second DNA construct comprising:
- i. a nucleic acid sequence encoding a heterologous polypeptide;
- ii. a m6A sensor sequence;
- and
- iii. a polypeptide encoding dihydrofolate reductase (DHFR).
4 more elements
A nucleic acid sequence comprising a nucleic acid sequence encoding a heterologous polypeptide, a m6A sensor sequence, and, a polypeptide encoding dihydrofolate reductase (DHFR).
A method for detecting in vitro m6A methylation in one or more cells comprising
- (a) contacting one or more cells with
- i. a fusion protein comprising an N6-methyladenosine (m6A) binding domain of a YT521-B homology (YTH) domain-containing protein fused to a catalytic domain of a cytidine deaminase or a catalytic domain of an adenosine deaminase;
- and
- ii. a DNA construct comprising a nucleic acid sequence encoding a heterologous polypeptide, a m6A sensor sequence, and a polypeptide encoding dihydrofolate reductase (DHFR);
- and
- (b) detecting expression of the reporter protein.
3 more elements
Dependencies are inferred from the claim text.