Towards epigenetics in sugar beet – the ONT based reference 5mC methylome of Beta vulgaris ssp. vulgaris

Epigenetic modifications, such as DNA methylation, influence phenotypic plasticity and affect numerous plant traits. Genome-wide DNA methylation patterns differ among cell types, species, and developmental stages. Still, it remains poorly understood in non-model plants. Thus, the potential of epigenetic breeding approaches targeting DNA methylation in crop plants is not fully realised. This study focuses on sugar beet ( Beta vulgaris L. ssp. vulgaris ) and comprises the first long read-based reference DNA methylome for this species, generated using Oxford Nanopore Technologies (ONT) sequencing. The detection of 5-methyl cytosine (5mC) in the three sequence contexts (CG, CHG, and CHH) was performed with DeepSignal-plant. The 5mCartograph tool was developed to provide a detailed overview of 5mC methylation probabilities: A genome-wide 5mC reference methylome was established for the genotype KWS2320, including gene- and repeat-specific analyses and the identification of 2,088 gene body methylated (gbM) genes, while 10,839 genes remained unmethylated. Genome-wide analysis re-detected more than 99% of the 204.8 Mio reference cytosines, based on ONT read sets of at least 17.5 × mapped genome coverage. Of these cytosines, 14.5% were classified as ‘highly methylated’ in young sugar beet leaves. Methylation levels followed typical plant patterns, being highest in CG (89.8%), followed by CHG (62.8%) and CHH (10.1%) context. This detailed methylome provides a robust foundation for future studies - such as epi-pangenome generation - and supports potential breeding applications in crop improvement. • ONT-based DNA methylation analysis delivers detailed insights into plant methylomes • Established generation of feature-specific methylation profiles by the 5mCartograph • DNA methylation patterns change along the chromosomes • DNA methylation patterns are highly feature specific in genes and repeats • Genome-wide DNA methylation patterns in sugar beet are context-specific