High-coverage allele-resolved single-cell DNA methylation profiling by scDEEP-mC reveals cell lineage, X-inactivation state, and replication dynamics

DNA methylation is a relatively stable epigenetic mark with important roles in development and disease.1 Since cell-to-cell variation in epigenetic programming can reflect essential differences in cell state and fate, it is clear that single-cell methods are necessary to strengthen our understanding of DNA methylation’s biological roles. Existing single-cell whole-genome bisulfite sequencing (scWGBS) methods2 have substantial shortcomings, including very low CpG coverage3–7 or inefficient library generation requiring extremely deep sequencing.8 We report an improved method, single-cell Deep and Efficient Epigenomic Profiling of methyl-C (scDEEP-mC). High-coverage DNA methylation profiling by scDEEP-mC allows cell type identification, genome-wide profiling of hemi-methylation, and allele-resolved analysis of X-inactivation epigenetics in single cells. We further demonstrate how scDEEP-mC can profile single-cell copy number alterations, identify replicating cells, and track DNA methylation dynamics during and after DNA replication. The high coverage of scDEEP-mC creates an exceptional opportunity to explore DNA methylation biology in individual cells. Single-cell Deep and Efficient Epigenomic Profiling of methyl-C (scDEEP-mC) was conducted on 175 single cells isolated from mouse intestinal epithelium, as well as 199 cultured human fibroblasts at different passage levels and 93 TERT-immortalized human fibroblasts.