DNA methylation analysis in plant gigagenomes: comparing two bisulfite sequencing techniques in Abies alba trees affected by dieback

Phenotypic plasticity is a fundamental mechanism that enables plants to adapt to shifting environmental conditions, such as those induced by climate change. Epigenetic modifications, notably DNA methylation, may play a pivotal role in such process. However, this field remains largely unstudied in non-model organisms with large, complex genomes. Here, we focus on silver fir (Abies alba), more precisely on a natural population subjected to climate stress, comparing the results obtained from two different bisulfite sequencing techniques in the study of the epigenetic patterns of its giga-genome. DNA was extracted from two non-declining and two declining A. alba individuals and subjected to whole genome bisulfite sequencing (WGBS) and reduced representation bisulfite sequencing (RRBS). General methylation levels in each cytosine context (CpG, CHG and CHH, where H stands for A, C, or T) were determined and differentially methylated regions (DMRs) were identified. WGBS sequenced a higher amount of cytosines, while RRBS had a higher number of CpG sites passing filtering conditions. Both techniques showed high methylation percentages in CpG and CHG contexts, but disagreed on CHH sites, likely due to RRBS having a bias towards highly methylated CHH contexts. Differentially methylated sites were only identified using WGBS data but could not be functionally annotated. In conclusion, while successful in methylation assessment, neither WGBS nor RRBS produced the desired outcome regarding the identification of DMRs. Thus, the fragmented nature of the available reference genomes for species with large genomes seems to hinder the attainment of meaningful results from WGBS and RRBS.