ORIGINAL PAPER Effect of liquid pulses with 6-benzyladenine on the induction of somatic embryogenesis from coffee (Coffea arabica L.) callus cultures Iosif Papanastasiou Æ Katerina Soukouli Æ Georgia Moschopoulou Æ Jane Kahia Æ Spiridon Kintzios Received: 5 November 2007 / Accepted: 25 November 2007 / Published online: 6 December 2007 Ó Springer Science+Business Media B.V. 2007 Abstract We investigated the effect of the physical state of the nutrient medium on the induction of somatic embryogenesis on cell cultures derived from coffee (Coffea arabica L.). Non-embryogenic callus tissues were pulsed initially with 50 lM 6-benzylad- enine (BA) for 6, 24 or 48 h in half-strength liquid Murashige and Skoog (MS) medium. After pretreat- ment, calli were transferred to agar-solidified half- strength MS medium supplemented with 50 lM BA (‘standard induction medium’). Control callus tissues were incubated directly on the solid standard induc- tion medium. Callus growth was promoted by longer pretreatment periods. Formation of globular somatic embryos was observed on callus tissues pretreated with BA for 24 or 48 h, which developed fully to cotyledonary-stage within only 2 weeks after transfer to agar-solidified medium supplemented with BA. No embryo formation occurred in control cultures. Pre- treatment with BA in liquid medium was associated with changes in the redox status of cultured cells, such as alterations of the ascorbate–glutathione redox systems and the accumulation of free radicals and oxidized lipids, as well as the possible reduction of cytochrome c-mediated apoptotic pathways. In par- ticular, the induction of somatic embryogenesis was highly positively correlated (r 2 = 0.822) with the accumulation of protein carbonyls. The physiological role of BA as an inducer of both embryonic differentiation and cellular death is discussed. Keywords Cell culture Á Embryo germination Á Leaf explant Á Indirect somatic embryogenesis Á Redox status Abbreviations AOX Ascorbate oxidase APX Ascorbate peroxidase Asc Ascorbate acid BA 6-Benzyladenine CDNB 1-Chloro-2,4-dinitrobenzene DHAR Deydroascorbate reductase DTT 1-4-Dithiothreitol GR Glutathione reductase GSSG Glutathione oxidised GSH Glutathione reduced HCl Hydrochloric acid HEPES N-2-Hydroxyethylpiperazine-N 0 -2- ethanesulfonic acid H 2 O 2 Hydrogen peroxide H 2 DCF-DA 2 0 ,7 0 -Dichlorofluorescein diacetate MDA Malondialdehyde MS Murashige and Skoog basal medium I. Papanastasiou Á K. Soukouli Á G. Moschopoulou Á S. Kintzios (&) Laboratory of Plant Physiology, Faculty of Agricultural Biotechnology, Agricultural University of Athens, Iera Odos 75, Athens 11855, Greece e-mail: skin@aua.gr J. Kahia International Coffee Research Institute, Ruiru, Kenya 123 Plant Cell Tiss Organ Cult (2008) 92:215–225 DOI 10.1007/s11240-007-9326-0