Efficient production of polyploid plants via protoplast culture of Iris fulva

Abstract

In protoplast culture of the Louisiana iris species, Iris fulva (2n=42), N6 medium supplemented with 1 mg/l 2,4-D, 1 mg/l kinetin, 200 mg/l casein hydrolysate, 250 mg/l proline, 0.3 M glucose and 2.5 g/l gellan gum was suitable for cell division and colony formation. When colonies formed were transferred to plant growth regulator free MS medium, 61 plant lines were regenerated via somatic embryogenesis. Flow cytometric analysis revealed that leaf tissue of I. fulva consisted of 2C, 4C and 6C cells. Consequently, we identified this species as polysomatic in the genus Iris. Such polysomaty was also recognized in the suspension calli. Sixty one plant lines obtained by protoplast culture consisted of 25 diploid (41.0%), 1 triploid (1.6%), 29 tetraploid (47.5%) and 6 hexaploid lines (9.9%). In addition, chromosome analysis showed that somatic chromosome numbers of the diploid, hypertriploid and tetraploid lines were 2n=42, 67 and 84, respectively. On the other hand, all lines regenerated from suspension culture of I. fulva were diploid lines in contrast to the results from protoplast culture. Thus, it can be concluded that protoplast culture of this species is an efficient technique for production of polyploid plants.