Induction and identification of polyploids in four Rhododendron species

To induce polyploidy in rhododendrons, different experimental materials of four Rhododendron species (2n = 2x = 26) were treated with colchicine and oryzalin as chemical mutagens. Three materials from R. fortunei, namely, seeds, cotyledons, and adventitious buds, were treated, and the highest tetraploid induction rate (15.64%) was observed in cotyledons treated with 20 mg·L^− 1 oryzalin for 4 d. Two materials (stem tips and stem bases) were treated in R. simsii, and the highest tetraploid induction rate (4.04%) was achieved with 0.14% colchicine after soaking the stem tips for 48 h. However, when we treated the stem bases, the chimerism occurrence rate was up to 30.00%. In addition, polyploidy induction in R. ovatum was carried out at two colchicine concentrations (0.12 and 0.14%) with two materials (stem tip and stem base), which resulted in a higher chimeric rate. Three materials (stem tip, stem base, and adventitious buds) of R. molle were used, but no polyploidy was found. Polyploid identification was performed through stomatal observation and flow cytometry. Polyploid plants had larger and sparser stomata than diploid plants. Furthermore, morphological measurements revealed that the size of the stomata and leaves in tetraploid plants was larger than that in diploid plants. Stomatal density was decreased in tetraploid plants compared to diploid plants. Flow cytometry analysis identified tetraploids in the mutant plants of R. fortunei and R. simsii. Phenotypic analyses showed that tetraploid plants had smaller, rounder leaves and darker leaf colors than diploid plants. In summary, we compared the induced polyploid rhododendrons using different methods, which is of great significance for cultivating new rhododendron varieties with good ornamental properties and high resistance and for developing the rhododendron industry in China.