(A and B) Analysis of nascent RNA synthesis in WT or Mettl3−/− mESCs [(A), Mettl3−/−−1 and −2 are two independently generated KO lines], and Mettl3−/− mESCs rescued with WT or an inactive mutant Mettl3 (B). Nascent RNA synthesis was detected by using a click-it RNA Alexa fluor 488 imaging kit. EU, 5-ethynyl uridine; DAPI, 4′,6-diamidino-2-phenylindole. (C and D) Analysis of chromatin accessibility in WT or Mettl3−/− mESCs (C), and Mettl3−/− mESCs rescued with WT or mutant Mettl3 (D). DNase I–treated TUNEL assay was performed. For (A) to (D), the nucleus is counterstained by DAPI. EV (empty vector) refers to Mettl3−/− mESCs when transfected with empty vector plasmid. dsDNA, double-stranded DNA.