N6-mA slows down UMP incorporation and promotes pol II backtracking. (a) The presence of N6-mA in the DNA template strand leads to slow UMP addition. The concentration of UTP is 25 μM. Time points are 0, 15 s, 30 s, 1 min, 2 min, 5 min, 20 min, and 1 h. (b) The presence of N6-mA in the DNA template promotes further backtracking revealed by TFIIS-stimulated transcript cleavage assay. Time points are 0, 15s, 1 min, 5 min, 20 min, 1 h, and 3 h, respectively. The DNA/RNA scaffolds are listed with i + 1 position highlighted. The n – 1 and n – 2 cleavage products are highlighted by black and blue arrows, respectively.