Response surface optimization of basal nutrients for in vitro conservation of endangered plants: a Gossia fragrantissima case study

Abstract

Standard commercially available basal media are commonly used in plant tissue culture but may not meet the specific nutritional needs of all species. This was a limiting factor for in vitro conservation of Gossia fragrantissima , an endangered Australian rainforest species threatened by myrtle rust, habitat loss, and grazing. To support its conservation, this study optimized the nutrient composition of the standard Woody Plant Medium (WPM) to enhance the physiological health of G. fragrantissima cultures for short-to-medium term conservation and as a pre-requisite for long-term cryopreservation. Using response surface methodology, six key nutrient factors (NH 4 NO 3 , Ca(NO 3 ) 2 , mesonutrients–CaCl 2 , KH 2 PO 4 , and MgSO 4 , micronutrients, Fe-EDTA, and WPM vitamins) were optimised. Shoot tip necrosis and defoliation were reduced by increasing Fe-EDTA and total nitrogen, while higher iron concentrations promoted shoot multiplication and elongation. An optimized medium was achieved by increasing NH 4 NO 3 to 2.5× WPM concentration, reducing Ca(NO 3 ) 2 to 0.5×, increasing mesonutrients to 2.37×, micronutrients to 4×, Fe-EDTA to 3.43×, and reducing WPM vitamins to 0.75×. This tailored formulation significantly improved growth and reduced undesirable traits compared to performance on WPM, demonstrating the critical role of customized basal nutrients in maintaining G. fragrantissima cultures. The findings provide a foundation for advancing tissue culture and cryopreservation protocols to ensure the long-term preservation of this endangered species.

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