Effects of various preculture, pretreatment, and recovery conditions on the morphogenetic response of cryopreserved Lady Orange chrysanthemum shoot tips

The aim of this research was to optimize the encapsulation-dehydration cryopreservation protocol of Lady Orange chrysanthemum explants on the preculture, pretreatment, and post-rewarming recovery steps. Shoot tips were precultured on MS medium with various abscisic acid concentrations (0-30 µM). Next, the encapsulated explants were osmotically dehydrated for 3 or 5 days and desiccated for 3-4 h and, after rewarming, they were subcultured on recovery media of various compositions (control and cytokinin/auxin-supplemented). A high explant survival rate, even up to 100%, was observed. The value of this parameter, however, changed depending on the post-rewarming culture duration. Moreover, not all the viable explants were capable of forming shoots. A lower ABA concentration (15 µM) during preculture and the presence of 4.65 µM kinetin in the post-rewarming recovery medium enhanced cryopreservation efficiency with a high survival rate and typical microshoot formation. A higher ABA concentration and the presence of 6-benzylaminopurine in the recovery medium resulted in shoot multiplication, abundant callus formation, and root formation inhibition.