Tetraploid Induction of Hibiscus moscheutos in Tissue Culture

Hibiscus moscheutos (2n=2x=38) is a perennial plant native to wetland areas in the eastern United States. Commercial breeding has focused on compact stature and enhanced floral characteristics. In recent years, interest has arisen to breed for purple foliage and sterility.  A purple-foliage selection (Hib14-113) was treated using colchicine and oryzalin as mitotic inhibitors to induce tetraploid breeding lines to be used for developing triploid genotypes. Hib14-113 plants were first transferred into tissue culture growing in MS basal media. After plants were well established in tissue culture, 10 two-node shoots per treatment were immersed in 0%, 0.025%, 0.05%, 0.1% (w/v) colchicine for 12, 24, 36 hours, and 0 μM, 100 μM, 125 μM, 150 μM oryzalin solutions for 3-, 6-, 9-hour durations on a rotational shaker (120 rpm). Following treatment the shoots were thoroughly rinsed with water and transferred back to MS basal media. Plants from each chemical treatment (n = 24) were placed under a 16-hour photoperiod and were arranged using a randomized complete block design with ten blocks. Survival of treated shoots was assessed six weeks after treatment. The ploidy level of plants was examined via flow cytometry. Following root establishment, plants were gradually acclimated to a greenhouse environment for evaluation of ornamental characteristics. After plants were determined to be diploid, mixaploid or tetraploid, they were all evaluated based on plant height, size, and other characteristics related to flowers, leaves and production of seeds. The concentration of colchicine (p<0.001) had an effect on survival of shoots. Colchicine had a higher lethal effect on the plants with 56.7% rate of survival, compared to 81.7% survival of oryzalin-treated plants. Both colchicine and oryzalin are considered effective polyploidy-inducing agents for Hibiscus moscheutos, and results on ploidy levels will be reported.