Development of NaCl-tolerant line in Chrysanthemum morifolium Ramat. through shoot organogenesis of selected callus line

Plants were regenerated successfully through shoot organogenesis of a NaCl-selected callus line of Chrysanthemum morifolium Ramat. cv. Maghi Yellow (a salt sensitive cultivar), developed through stepwise increase in NaCl concentration (0–100 mM) in the MS medium. The stepwise increase in NaCl concentration from a relatively low level to cytotoxic level was found to be a better way to isolate NaCl-tolerant callus line, since direct transfer of callus to high saline medium was detrimental to callus survival and growth. The selected callus line exhibited significant increase in superoxide dismutase (EC 1.15.1.1), ascorbate peroxidase (EC 1.11.1.11) and glutathione reductase (EC 1.6.4.2) activities compared to control callus (grown in medium devoid of NaCl). Stability of salt tolerance character of the selected callus line was checked by growing the calli in NaCl-free medium for 3 consecutive months followed by re-exposure to higher salinity stress (120 mM NaCl). Among different growth regulator treatments, a combination of 5 mg l−1 TDZ (Thidiazuron) along with 0.25 mg l−1 NAA and 0.5 mg l−1 GA3 was found to be the most effective for shoot organogenesis in selected callus line. The regeneration potential of the NaCl-tolerant callus ranged from 20.8% to 0% against 62.4% to 0% in control callus line. Under elevated stress condition (medium supplemented with 250 mM NaCl), selected calli derived regenerants (S1 plants) exhibited significantly higher SOD and APX activities over both PC (positive control: control callus derived plants grown on MS medium devoid of NaCl) and NC (negative control: control callus derived plants subjected to 250 mM NaCl stress) plants. In addition, the NC plants showed stunted growth, delayed root initiation, and had lesser number of roots as compared to S1 plants. Based on growth performance and antioxidant capacity, the S1 plants could be considered as NaCl-tolerant line showing all positive adaptive features towards the salinity stress. Further study on agronomic performance of these S1 plants under saline soil condition need to be undertaken to check the genetic stability of the induced salt-tolerance.