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Background  As an important epigenetic mark, 5-methylcytosine (5mC) methylation is involved in many DNA-dependent biological processes and plays a role during development and differentiation of multicellular organisms. However, there is still a lack of knowledge about the dynamic aspects and the roles of global 5mC methylation in wood formation in tree trunks. In this study, we not only scrutinized single-base resolution methylomes of primary stems (PS), transitional stems (TS), and secondary stems (SS) of  Populus trichocarpa  using a high-throughput bisulfite sequencing technique, but also analyzed the effects of 5mC methylation on the expression of genes involved in wood formation.    Results  The overall average percentages of CG, CHG, and CHH methylation in poplar stems were ~ 53.6%, ~ 37.7%, and ~ 8.5%, respectively, and the differences of 5mC in genome-wide CG/CHG/CHH contexts among PS, TS, and SS were statistically significant ( p  < 0.05). The evident differences in CG, CHG, and CHH methylation contexts among 2 kb proximal promoters, gene bodies, and 2 kb downstream regions were observed among PS, TS, and SS. Further analysis revealed a perceptible global correlation between 5mC methylation levels of gene bodies and transcript levels but failed to reveal a correlation between 5mC methylation levels of proximal promoter regions and transcript levels. We identified 653 and 858 DMGs and 4978 and 4780 DEGs in PS vs TS and TS vs SS comparisons, respectively. Only 113 genes of 653 DMGs and 4978 DEGs, and 114 genes of 858 DMGs and 4780 DEG were common. Counterparts of some of these common genes in other species, including  Arabidopsis thaliana , are known to be involved in secondary cell wall biosynthesis and hormone signaling. This indicates that methylation may directly modulate wood formation genes and indirectly attune hormone signaling genes, which in turn impact wood formation.    Conclusions  DNA methylation only marginally affects pathway genes or regulators involved in wood formation, suggesting that further studies of wood formation should lean towards the indirect effects of methylation. The information and data we provide here will be instrumental for understanding the roles of methylation in wood formation in tree species.

DNA methylation and its effects on gene expression during primary to secondary growth in poplar stems