Sequencing of N6-methyl-deoxyadenosine at single-base resolution across the mammalian genome

Mol Cell. 2024 Feb 1;84(3):596-610.e6. doi: 10.1016/j.molcel.2023.12.021. Epub 2024 Jan 11.


Although DNA N6-methyl-deoxyadenosine (6mA) is abundant in bacteria and protists, its presence and function in mammalian genomes have been less clear. We present Direct-Read 6mA sequencing (DR-6mA-seq), an antibody-independent method, to measure 6mA at base resolution. DR-6mA-seq employs a unique mutation-based strategy to reveal 6mA sites as misincorporation signatures without any chemical or enzymatic modulation of 6mA. We validated DR-6mA-seq through the successful mapping of the well-characterized G(6mA)TC motif in the E. coli DNA. As expected, when applying DR-6mA-seq to mammalian systems, we found that genomic DNA (gDNA) 6mA abundance is generally low in most mammalian tissues and cells; however, we did observe distinct gDNA 6mA sites in mouse testis and glioblastoma cells. DR-6mA-seq provides an enabling tool to detect 6mA at single-base resolution for a comprehensive understanding of DNA 6mA in eukaryotes.

Keywords: DNA modification; N(6)-methyl-deoxyadenosine; epigenetics; glioblastoma; single-base resolution sequencing.

MeSH terms

  • Animals
  • DNA / metabolism
  • DNA Methylation*
  • Deoxyadenosines / genetics
  • Escherichia coli* / genetics
  • Eukaryota / genetics
  • Genome / genetics
  • Mammals / metabolism
  • Mice


  • DNA
  • Deoxyadenosines