Objective Four polyploid germplasm of Iris germanica were studied to facilitate resistance improvement and value enhancement of the ornamental plants.

Method Tissue culture clusters collected from I. germanica varieties, Castle, Black Knight, Black Mercury, and Cimalen Zone, were cultured on the clump-forming bud induction medium and rooting medium containing varied concentrations of colchicine (COL) to grow mutant polyploid plants. Ploidy of the plants were identified morphologically, cytologically, and molecular biologically.

Result The optimal medium for inducing the mutant Castle bud clusters was formulated with MS + 0.5 mg·L⁻¹ 6-BA + 0.5 mg·L⁻¹ NAA + 0.10% COL, and MS + 0.5 mg·L⁻¹ 6-BA + 1.0 mg·L⁻¹ NAA + 0.10% COL for Black Mercury. The optimized rooting medium for both was same with 1/2MS + 0.5 mg·L⁻¹ NAA + 0.10% COL. They resulted in a mutation induction rate of 80.00±0.37% on Castle and 66.67±0.12% on Black Mercury. The selected induction medium for Black Knight contained MS + 0.5 mg·L⁻¹ 6-BA + 0.5 mg·L⁻¹ NAA + 0.05% COL, and that for Cimalen Zone MS + 1.5 mg·L⁻¹ 6-BA + 1.0 mg·L⁻¹ NAA + 0.05% COL. While the identical rooting medium for the two consisted of 1/2MS + 0.5 mg·L⁻¹ NAA + 0.05% COL. The final induction rate was 46.67±0.08% for Black Knight, and 73.33±4.22% for Cimalen Zone. Morphologically, all mutant plants showed polyploid characteristics such as greater height, wider and thicker leaves, stronger roots, larger stomata, and lower stomatal density than the comparative regular plants. The chromosome counts at root tips of the 4 mutant germplasms were twice that of control, and the ploidy identified by flow cytometry with DNA fluorescence intensity twice that of non-mutant originals as well. Thus, the chromosome doubling plants were successfully generated.

Conclusion The ploidy of the mutant Castle and Black Mercury were both 2n = 64 (versus 2n = 32 of control), that of mutant Black Knight 2n = 48 (vs. 2n = 24 of control), and that of mutant Cimalen Zone 2n = 44 (vs. 2n = 22 of control). The system developed in this study that satisfactorily created 4 polyploid I. germanica germplasms seemed to be technically applicable for generating new varieties to meet a broadened landscaping demand.