Whole Genome Bisulfite Sequencing in wild type plants, and two RNAi transgenic lines, KD2 and KD5.. Populus tremula x Populus alba

We report the application of Whole Genome Bisulfite Sequencing to identify differentially methylated regions (DMR) in two RNAi transgenic lines, with dowregulated expression of the demeter-like gene PtaDML10, in comparisson to wild-type plants of Populus tremula x Populus alba plants. Overall design: wild type and two PtaDML10 RNAi lines, KD2 and KD5, were transferred to 3.5 L pots containing blond peat, pH 4.5. Before any photoperiodic treatments, the plants were grown in a chamber under LD controlled conditions (16h light, 21°C, 65% relative humidity and 150 µmol m-2 s-1 light intensity). After 4 weeks the plants were transferred to SD conditions (8h light, 19°C, 65% relative humidity and 150 µmol m-2 s-1 light intensity) for 12 weeks in order to induce apical bud formation. To mimic winter conditions, dormant plants were kept under SD, 100 µmol m-2 s-1 light intensity and 4°C for 4 weeks. The topmost bud of each plant, so-called apical, was collected when the first visual changes in bud developmental were evident. Buds with morphological changes indicating initial stages of bud break were removed, so all the apical buds were collected in stage 0. We collected 15 apical buds from 15 plants per each WT, KD2 and KD5 genotypes for whole genome bisulfite sequencing analyses.