In vitro induction of tetraploid Ziziphus jujuba Mill. var. spinosa plants from leaf explants

Tetraploid Ziziphus jujuba Mill. var. spinosa plants were induced by combining the adventitious shoot organogenesis from leaf explants and polyploidization with colchicine. Leaf explants were cultured on WPM containing1.0 mg L−1 thidiazuron and 0.3 mg L−1 3-indolyl-butyric acid for 10 or 20 days. They were then transferred to the same liquid medium supplemented with 40, 60, or 80 mg L−1 colchicine and incubated for 24, 48, or 72 h. The highest tetraploid induction rate (6.67%) resulted from a 10-day pre-culture treatment followed by an incubation in 60 mg L−1 colchicine for 72 h. The ploidy levels of the regenerated shootlets were determined based on flow cytometry analysis and chromosome numbers. Additionally, there were significant morphological differences between the tetraploid and diploid plants regarding stomatal size and density as well as leaf and shoot characteristics. This variability represents an early screening to distinguish the shoot with a ploidy level alternation from the diploid ones. The initial field performance results showed a faster growth in tetraploids than in the diploids, as suggested by the basal diameter and plant height increases.