Abiotic stressors such as drought, salinity, and exposure to heavy metals can induce epigenetic changes in plants. In this study, liquid chromatography (RP-HPLC), methylation amplified fragment length polymorphisms (metAFLP), and methylation-sensitive amplification polymorphisms (MSAP) analysis was used to investigate the effects of aluminum (Al) stress on DNA methylation levels in the crop species triticale. RP-HPLC, but not metAFLP or MSAP, revealed significant differences in methylation between Al-tolerant (T) and non-tolerant (NT) triticale lines. The direction of methylation change was dependent on phenotype and organ. Al treatment increased the level of global DNA methylation in roots of T lines by approximately 0.6%, whereas demethylation of approximately 1.0% was observed in NT lines. DNA methylation in leaves was not affected by Al stress. The metAFLP and MSAP approaches identified DNA alterations induced by Al3+ treatment. The metAFLP technique revealed sequence changes in roots of all analyzed triticale lines and few mutations in leaves. MSAP showed that demethylation of CCGG sites reached approximately 3.97% and 3.75% for T and NT lines, respectively, and was more abundant than de novo methylation, which was observed only in two tolerant lines affected by Al stress. Three of the MSAP fragments showed similarity to genes involved in abiotic stress.
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