Assessing different storage temperatures and protocols for preserving seeds and nodal explants of the African leafy vegetable, Amaranthus dubius

Abstract

This study investigated the response of seeds and nodal explants of the highly nutritious African leafy vegetable (ALV), Amaranthus dubius, to multiple cryopreservation protocols (controlled-rate cooling, liquid nitrogen (LN) immersion, and LN slurry (semi-solid mix of liquid and frozen nitrogen) immersion) and storage temperatures (24 (room temperature), 4 (fridge), -20 (freezer), -80 (ultrafreezer), -148 (LN vapour) and -196 °C (LN)). This study revealed that A. dubius seeds can be stored for 4 years at -20 and -80 °C without significantly decreasing germination capacity (86 and 88 %, respectively) compared to the non-stored control (92%). Cooling seeds at a controlled rate was found to be optimal in maintaining seed viability, irrespective of storage temperature. Controlled-rate cooling was also optimal for nodal explants across storage temperatures (-20, -80, -148, and -196 °C), which, when multiplied in vitro, yielded 3.6 ± 0.6 shoots per explant with an 80 % shoot development rate. To assess the effect of these treatments on genetic fidelity, a previously identified salinity tolerant A. dubius genotype, expressing a Na+/H+ exchanger (NHX1) gene, was stored, and the relative expression of the NHX1 transcript was quantified in root tissues post-acclimatisation. Nodes that were cooled at a controlled rate, in LN slurry, or in LN, maintained genetic fidelity better than those of the control, based on the relative expression of the NHX1 marker gene.