Seed sterilisation and germination

Timing ~1 h (for sterilisation, depending on the amount of seeds), 6 days (for germination)

Sterilisation solution should be prepared freshly. Seeds are incubated for 2 min in 2 ml 100 % ethanol and subsequently surface sterilised with 2 ml sterilisation solution for 15 min and then washed with sterile water three times for 10 min [37]. A better washing can be achieved by shaking the reaction tube containing the seeds.

Transfer the sterilised seeds to the germination plate (6–8 seeds per plate) and incubate the plates at 4 °C in the dark for 3 days in a vertical orientation.

After 3 days incubate the plates under short day conditions (light: 8 h, dark: 16 h, light intensity 100 μmol m⁻² s⁻¹, 22 °C and a relative humidity of 70 %) in a growth chamber and store the plates in a vertical orientation. After approx. 6 days old 3–5 cm long seedlings were used for grafting.