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Adventitious shoot regeneration of two dwarfing pear rootstocks and the development of a transformation protocol

Summary The ability to form adventitious shoots was investigated from in vitro produced leaves of the dwarfing pear rootstock BP10030 and the semi-dwarfing pear rootstock OHF333. To increase rooting ability, transformation of the rootstock BP10030 with the rolB gene was performed. For BP10030, 98% leaves gave rise to shoots on the medium containing 1 μM NAA, 15 μM TDZ and sucrose, and 88% from the medium with 0.1 μM NAA, 5 μM TDZ and sorbitol. For OHF333, the best result was obtained from the medium containing 0.1 μM NAA, 1 μM TDZ and sorbitol, using which 66% of leaves gave rise to shoots, whereas the regeneration percentage was generally low on the sucrose medium (0–22%). In the experiments where IBA was used as a substitute for NAA and CPPU as a substitute for TDZ on the sucrose medium, shoot regeneration of OHF333 was not improved. Scanning electron microscopy (SEM) observations on BP10030 revealed that meristem-like structures were already visible at day 8 after the beginning of the experiments and well-developed shoots appeared after one month. SEM also showed that shoots arose from the surface of the callus and more meristems were observed than shoots eventually formed. Transformation experiments on BP10030 using the published protocol failed. However, transformation based on the results from the regeneration tests was successful, indicating genotype differences and that a good protocol for adventitious shoot regeneration is necessary for a successful transformation. However, the transformation efficiency was low, suggesting that a good regeneration protocol cannot guarantee high transformation efficiency.