Growth and physiological characteristics of wasabi plantlets cultured by photoautotrophic micropropagation at different temperatures

Iranian seedless barberry is a very recalcitrant species in in vitro culture which does not show appropriate growth on standard culture media. Response surface methodology was employed to evaluate the effects of changing macronutrients concentrations on establishment and proliferation phases. KNO₃ and NH₄NO₃ macronutrients at 0.3 to 1.5?×?MS medium levels and CaCl₂, MgSO₄ and KH₂PO₄ macronutrients in a range of 0.5 to 1.5?×?MS medium concentrations were tested in a response surface design with 30 treatments. Many significant interactions were found among the macronutrients. High concentrations of KNO₃, NH₄NO₃ and CaCl₂ improved the growth rate in the establishment phase. The growth rate in media containing high KNO₃ and low CaCl₂ was high. Reduced concentrations of CaCl₂ and KNO₃ decreased hyperhydricity. The greatest hyperhydricity was induced when both NH₄NO₃ and CaCl₂ were used at 1.5?×?MS level. The number of hooked leaves decreased as KH₂PO₄ increased and MgSO₄ reduced. In the proliferation phase, there were many significant interactions among the macronutrients. Increased concentration of NH₄NO₃ and reduced concentration of KH₂PO₄ improved the growth rate. Proliferation rate increased in media containing high concentration of KNO₃ and low to moderate concentrations of NH₄NO₃. The greatest production of new tissues and organs was seen in media with high KNO₃ and moderate to high CaCl₂. High concentration of NH₄NO₃ and low concentration of KH₂PO₄ also increased production of new tissues and organs. No shoot apical meristem was seen when CaCl₂ level was high and KNO₃ level was low. Formation of shoot apical meristem required high KH₂PO₄ concentration and low CaCl₂ concentration. Finally, low concentration of KNO₃ and low to moderate concentrations of NH₄NO₃ increased phenol exudation.