Pre-culture before Agrobacterium Infection to Leaf Segments and Meropenem Improves the Transformation Efficiency of Apple (Malus * domestica Borkh.)

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Transformation experiments in apple (Malus × domestica Borkh.) were conducted to improve the Agrobacterium-mediated transformation efficiency of many apple cultivars without using complicated procedures. A system of direct regeneration from leaf segments was used for these experiments. The effectiveness of meropenem hydrate (MEPM) as an antibiotic for Agrobacterium elimination was investigated for use of instead of cefotaxime sodium (CTX), which prevents shoot regeneration from apple leaf segments. Results show that 50 mg·L−1 of MEPM greatly enhanced the transformation efficiency. Experiments assessing the timing of kanamycin sulfate (KM) addition to the medium were also conducted to investigate the effective use of KM, the antibiotic used to select transformed cell. Addition of KM to the medium immediately after co-cultivation was completed is advantageous for the multiplication of transformed cells and for the induction of transformed shoots in spite of the low shoot regeneration rate. Based on knowledge of the timing of KM addition, the effects of pre-culture before Agrobacterium infection on the shoot regeneration medium were investigated. Pre-culture for 3–7 days improved the transformation efficiency. By applying both MEPM and the pre-culture, about 5% transformation efficiency in 'Greensleeves' was achieved on an apple cultivar without complicated procedures.
園芸学会の論文