Evaluation of total protein, peroxidase, and nutrients measured by pXRF for the determination of tissue rejuvenation / reinvigoration of Eucalyptus microcorys

Ontogenetic aging of tissues and the gradual decrease of adventitious rooting are known challenges for the clonal propagation of woody species, hampering clonal forestry programs. This study examined possible signatures of tissue rejuvenation/reinvigoration in different propagated materials of Eucalyptus microcorys by analyzing the total protein profile, peroxidase activity, macro-and micronutrient contents, and adventitious rooting of mini cuttings. The analyses were performed on E. microcorys shoots which were successfully obtained by seminal and grafting propagation, micropropagation with epicormic shoots, and indirect organogenesis. Among four mature trees used in the propagation, tissues from the one with the best propagation results were investigated for signs of tissue rejuvenation and/or reinvigoration. Five individuals from each technique were randomly selected and transferred to a semi-hydroponic “channel” system. After four weeks in the seedbed, the total protein, peroxidase activity, nutrient content and rooting of the minicuttings were evaluated. SDS-PAGE enabled the differentiation of leaf samples obtained by grafting from the other propagation techniques, as revealed by two distinct bands. Materials obtained by micropropagation with epicormic shoots showed the highest peroxidase activity, while those obtained by seminal propagation and from the selected mature tree showed the lowest peroxidase activity. A portable X-ray fluorescence spectroscope (pXRF) identified adequate nutrient content in most of the nutrients tested in materials obtained by seminal and grafting propagation, and by indirect organogenesis. The analysis of adventitious rooting showed that the highest rooting percentage was observed in mini cuttings from seminal propagation (75%) followed by indirect organogenesis (35%). Based on principal component analysis, it was concluded that rooting of mini cuttings from both seminal propagation and indirect organogenesis organogenesis was associated with phosphorous, sulphur, and potassium contents, which suggests a higher level of tissue rejuvenation/reinvigoration in these propagated plants. Further studies are recommended to search for other methods that present similarities with the responses to adventitious rooting in forest species and thus optimize the rescue and propagation of plants with distinct ontogenetic stages.