Metabolic incorporation of bioorthogonal functional groups into cellular nucleic acids can be impeded by insufficient phosphorylation of nucleosides. Previous studies found that 5azidomethyl‐2′‐deoxyuridine (AmdU) was incorporated into the DNA of HeLa cells expressing a low‐fidelity thymidine kinase, but not by wild‐type HeLa cells. Here we report that membrane‐permeable phosphotriester derivatives of AmdU can exhibit enhanced incorporation into the DNA of wild‐type cells and animals. AmdU monophosphate derivatives bearing either 5′‐bispivaloyloxymethyl (POM), 5′‐bis‐(4‐acetoxybenzyl) (AB), or “Protide” protective groups were used to mask the phosphate group of AmdU prior to its entry into cells. The POM derivative “POM‐AmdU” exhibited better chemical stability, greater metabolic incorporation efficiency, and lower toxicity than “AB‐AmdU”. Remarkably, the addition of POM‐AmdU to the water of zebrafish larvae enabled the biosynthesis of azide‐modified DNA throughout the body.