CUT&Tag-BS for simultaneous profiling of histone modification and DNA methylation with high efficiency and low cost.

It remains a challenge to decipher the complex relationship between DNA methylation, histone modification, and the underlying DNA sequence with limited input material. Here, we developed an efficient, low-input, and low-cost method for the simultaneous profiling of genomic localization of histone modification and methylation status of the underlying DNA at single-base resolution from the same cells in a single experiment by integrating cleavage under targets and tagmentation (CUT&Tag) with tagmentation-based bisulfite sequencing (CUT&Tag-BS). We demonstrated the validity of our method using representative histone modifications of euchromatin and constitutive and facultative heterochromatin (H3K4me1, H3K9me3, and H3K27me3, respectively). Similar histone modification enrichment patterns were observed in CUT&Tag-BS compared with non-bisulfite-treated control, and H3K4me1-marked regions were found to mostly be CpG poor, lack methylation concordance, and exhibit prevalent DNA methylation heterogeneity among mouse embryonic stem cells (mESCs). We anticipate that CUT&Tag-BS will be widely applied to directly address the genomic relationship between DNA methylation and histone modification, especially in low-input scenarios with precious biological samples.