DNA methylation assay of the RhAG promoter by bisulfite sequencing. Genomic DNA was isolated from flower buds grown under normal temperature (25/15 °C) or low temperature (15/5 °C) conditions. Bisulfite-converted DNA was amplified and sequenced. The sequences were analyzed using the CyMATE programme . The 166 bp DNA sequence analyzed by bisulfite sequencing is presented as top diagram. The number on the top indicate the base pair in the sequence; the number on the bottom indicate the cytosines in the sequence. Green triangles, blue squares and red circles represent cytosines in CHH, CHG and CG (H represents A, T, or C) configurations, respectively. Filled shapes indicate methylated sites, while open shapes indicate sites that are not methylated. Seventeen clones were analyzed for each treatment. The cytosines density is indicated by connection lines at the top panel as described in Hetzl et al.