An Optimized Chemical-Genetic Method for Cell-Specific Metabolic Labeling of RNA. An Optimized Chemical-Genetic Method for Cell-Specific Metabolic Labeling of RNA

We demonstrate that mammalian cells are able to salvage uracil analogs, yielding high background incorporation into cellular RNA. We also discover the enzymatic pathways responsible for this background. To overcome these limitations, we have developed a novel small-molecule/enzyme pair consisting of uridine/cytidine kinase 2 (UCK2) and 2'-azidouridine (2'AzUd). Overall design: Six total samples, consisting of three biological replicates each of HEK293T-UCK2/NIH3T3 co-culture treated with 2'AzUd 1 mM for 12 h Input and HEK293T-UCK2/NIH3T3 co-culture treated with 2'AzUd 1 mM for 12 h Enriched