Cytokinin oxidase/dehydrogenase inhibitors stimulate 2iP to induce direct somatic embryogenesis in Coffea arabica

Somatic embryogenesis in Coffea spp. is important for mass production and genetic engineering. Coffee is an exceptional case, as somatic embryogenesis can be induced by applying a cytokinin such as 2iP as the only plant growth regulator. 1-(2-(2-Hydroxyethyl)phenyl)-3-(3-(trifluoromethoxy)phenyl)urea (3TFM-2HE) and 1-(3-bromo-5-(trifluoromethoxy)phenyl)-3-(2-(2-hydroxyethyl)phenyl)urea (3TFM,5Br-2HE) are two newly designed cytokinin oxidase/dehydrogenase inhibitors (CKX) from the diphenylurea group. We used a Coffea arabica leaf disks bioassay to demonstrate the indirect somatic embryo induction potential of these compounds. The leaf disks were incubated on modified Murashige and Skoog (MS) semi-solid medium in which 3TFM-2HE or 3TFM,5Br-2HE were combined with N-6-(2-isopentenyl)adenine (2iP). Although these compounds do not possess intrinsic cytokinin activity, they enhanced the activity of 2iP, resulting in direct somatic embryogenesis after seven weeks. The best results were obtained with 1 mu M 3TFM-2HE and 5 or 10 mu M 2iP. Maturation of somatic embryos into fully developed plants took place on medium supplemented with 0.5 mu M kinetin and the somatic embryos developed true leaves and a root system.