In vitro-induced tetraploidy in sugarcane (Saccharum officinarum L. cv. CP57) using colchicine: protocol optimization and cytomorphological characterization

An efficient in vitro protocol was developed for inducing stable tetraploidy in sugarcane ( Saccharum officinarum L. cv. CP57), a cultivar with restricted flowering that hinders conventional breeding. In vitro -grown plantlets were treated with colchicine (0, 50, 100, and 200 mg L⁻¹) for 24, 48, or 72 h. The 72 h treatments resulted in complete mortality. The optimal treatment for polyploidy induction was 50 mg L⁻¹ colchicine for 24 h, which yielded the highest survival rate (83.3 ± 4.4%) and successfully produced tetraploids. Morpho-physiological analysis 60 days post-acclimatization revealed that this treatment significantly reduced tiller number (by 58%) and plant height (by 18%) while increasing chlorophyll content (by 18%) compared to diploid controls. Stomatal density decreased by 76%, while stomatal length and width increased by 150% and 120%, respectively, serving as reliable indicators of polyploidy. Cytological analysis confirmed tetraploidy (2n = ~230 chromosomes) in 68% of cells from plantlets treated with 50 mg L⁻¹ colchicine for 24 h. Karyotype analysis showed a 58% increase in total chromosome length and enhanced karyotype symmetry in tetraploids. The established protocol effectively induces tetraploidy in sugarcane cv. CP57 and provides a valuable biotechnological tool for breeding programs aimed at enhancing genetic diversity and improving agronomic traits.