Shoot organogenesis and somatic embryogenesis from leaf and petiole explants of endangered Euryodendron excelsum

Abstract

Euryodendron excelsum H.T. Chang is a first class of rare and endangered woody plant endemic to China. Leaves and petioles from the axillary shoots in vitro were used as explants to culture on the different PGR WPM medium and establish an efficient shoot proliferation and plant regeneration system. Callus and 3–6 adventitious shoots or somatic embryos were induced. WPM supplemented with 1.0 mg/L 2,4-D induced callus that then dedifferentiated into shoots and somatic embryos on different media, including PGR-free WPM. However, only adventitious shoots formed on WPM with 1.0 mg/L of cytokinins, 6-benzyladenine (BA), kinetin (KIN) or thidiazuron (TDZ). When another cytokinin, zeatin, was used, somatic embryos were induced directly from leaves and petioles. Adventitious roots could be induced from both explants on WPM with 1.0 mg/L α-naphthaleneacetic acid (NAA). Somatic embryos cultured in PGR-free WPM or WPM with 0.2 mg/L NAA developed roots. Plantlets derived from somatic embryos were transferred to a peat: sand (1:1, v/v) substrate, with 64.3% survival after 30 d, and 54.6% after 90 d. Callus clumps with adventitious shoot buds that were transferred to WPM with 1.0 mg/L BA and 0.2 mg/L NAA developed mean 3.3 multiple shoots. Callus-derived shoots regenerated and rooted successfully (100%) on agar-free vermiculite-based WPM with 0.5 µM NAA after 30 d. Plantlets transplanted to peat soil: vermiculite (1:1, v/v) displayed highest survival (96.7%) after three months. This protocol allows for the mass propagation of tissue-cultured plants and the conservation of this rare and endangered genetic resource.