A protocol for the micropropagation of Eucalyptus polybractea R.T. Baker (blue mallee) using axillary bud proliferation from lignotuber-derived explants is described. Three different ages of plants were used as explant sources: glasshouse-grown seedlings, field-grown saplings, and coppice of field-grown mature lignotubers. Explants from each source initiated successfully and no significant difference was observed for shoot proliferation, rooting success or hardening success between explant sources. Leaf oil quantity and quality for hardened clones transplanted to a field plantation were assessed after 3 months of growth. Ramets of all clones contained high quality oil with over 80% 1,8-cineole. For seedling-derived clones, foliar oil concentrations of ramets were higher than those of the ortets from which they were derived. For sapling and mature lignotuber derived clones the opposite was the case. This suggests that ontogenetic and physiological constraints may be influencing yield in the young ramets. The age of the explant source did not appear to influence the success of micropropagation, and as a result older plants (for which key oil traits are known) can be selected as elite plants for multiplying selected genotypes via micropropagation.