Resumen
Aim of study: The aim of the work was to determine the optimum axillary shoot induction system by optimizing different factors.Area of study: The different factors which were pretreatment methods, media types, kinds of cytokinins and dark treatmentMaterial and methods: After pretreating and sterilizing, the stems of Wollemi pine which about 3cm long were cultured on three kinds of media (GD, DCR AND ms) with different concentration and type of cytokines (BA, ZT and TDZ), as well as dark treatment (0, and 4 weeks) and then exposure to tissue culture room. The experiments were randomized block designed and the results were analyzed with ANOVA.Main results: Rapid freezing in liquid nitrogen was the most effective pretreatment method, dark treatment for the first 4-weeks of stem explants cultured in induction media could increase the frequency of axillary shoot induction significantly. The highest frequency of axillary shoot induction (63%) was obtained when explants were cultured in 1/2GD medium with 6.8 µM TDZ combining 4-weeks dark treatment. Research highlights: The optimized conditions for shoot production in vitro can be useful for conservation in vitro of Wollemia nobilis.Keywords: Wollemia nobilis; in vitro conservation; tissue culture; an endangered relict conifer.