Fig. 3

6mA directs nucleosome positioning in vitro. a Scheme of in vitro nucleosome assembly on native genomic DNA. Histone H2A, H2B, H3, and H4 are represented as different color balls. DNA is represented as dark curves. The red stars on DNA represent 6mA modification. Scissors represent endonuclease MNase which preferentially digests linker DNA. b The sequencing profile of nucleosomes assembled on native genomic DNA. Nucleosome centers were predicted and aligned to the flanking 2 kbp region of each TSS, and the accumulative occupancy was calculated and plotted around TSS. c Scheme of in vitro nucleosome assembly on unmethylated genomic DNA. Unmethylated DNA was acquired from whole genome amplification (WGA) from 5 ng genomic DNA. d The sequencing profile of nucleosomes assembled on unmethylated genomic DNA from WGA. Nucleosome centers were predicted and aligned to the flanking 2 kbp region of each TSS, and the accumulative occupancy was calculated and plotted around TSS. e Comparison of nucleosome distributions around single 6mA sites from three independent conditions: in vivo, native nucleosome profile; in vitro, nucleosome assembly on genomic DNA; and in vitro (UM), nucleosome assembly on unmethylated DNA. f Scheme of in vitro nucleosome assembly on model DNA. Unmethylated DNA was mixed with Dam-treated DNA at 1:1 ratio. g 6mA level of nucleosome-protected model DNA versus input DNA and unprotected regions. After in vitro nucleosome assembly followed by MNase digestion, UHPLC-QQQ-MS/MS was used to measure the 6mA abundance of input DNA, DNA regions resisting MNase digestion (nucleosome protected), and digested DNA (flowthrough). Error bars indicate mean ± s.d. of three technical replicates, each measured in duplicates. **p < 0.01 by Student’s t test