Analysis of nutrient deficiencies affecting in vitro growth and development of Eucalyptus dunnii Maiden

Although basal medium optimization is a key factor in the success of tissue culture, its mineral composition is frequently disregarded when optimizing in vitro propagation protocols. A previous work on Eucalyptus dunnii micropropagation suggests that excessive callus formation and leaf chlorosis are related to specific nutritional conditions of the basal media. Recently, a novel basal medium based on the mineral nutrient analysis of E. dunnii young stump shoots was developed and successfully tested in plant regeneration and micropropagation of E. dunnii , avoiding all these issues. Considering this basal medium as an ideal growth condition, a mild deprivation of each macro and micronutrient and of the total organic fraction was imposed to E. dunnii in vitro cultures for 30 d. As a result, K, Mg, Mn, Cl, Zn, Mo, Ni or Co deprivation quantitatively affected growth and development of axillary shoots. Moreover, leaf chlorosis and the development of organogenic callus under Fe deficiency, and leaf drop along with shoot tip necrosis under N deficiency were observed. These symptoms suggest that nutrient content in E. dunnii tissues needs to be above 420.3 mg kg −1 for Fe and 27.7 g kg −1 for N to avoid the symptoms of leaf chlorosis and shoot tip necrosis. Additionally, the main role of Mn in quantitative responses and the antagonism between ions, especially for Mg/K and Mg/Zn, were denoted by the multivariate analysis. Overall, these results make a relevant contribution to the optimization of in vitro propagation of E. dunnii and other hard-to-propagate related species.