AGROBACTERIUM-MEDIATED TRANSFORMATION OF APPLE (MALUS X DOMESTICA BORKH) - AN ASSESSMENT OF FACTORS AFFECTING GENE-TRANSFER EFFICIENCY DURING EARLY TRANSFORMATION STEPS

The factors influencing transfer of an intron - containing beta-glucur onidase gene to apple leaf explants were studied during early steps of an Agrobacterium tumefaciens-mediated transformation procedure, The g ene transfer process was evaluated by counting the number of beta-gluc uronidase expressing leaf zones immediately after cocultivation, as we ll as by counting the number of beta-glucuronidase expressing calli de veloping on the explants after 6 weeks of postcultivation in the prese nce of 50 mg/l kanamycin. Of three different tested disarmed A. tumefa ciens strains, EHA101(pEHA101) was the most effective for apple transf ormation. Cocultivation of leaf explants with A. tumefaciens on a medi um with a high cytokinin level was more conducive to gene transfer tha n cocultivation on media with high auxin concentrations, Precultivatio n of leaf explants, prior to cocultivation, slightly increased the num ber of beta-glucuronidase expressing zones measured immediately after cocultivation, but it drastically decreased the number of transformed calli appearing on the explants 6 weeks after infection. Other factors examined were: Agrobacterium cell density during infection, bacterial growth phase, nature of the carbon source, explant age, and explant g enotype.