Establishment of an in vitro Culture and Regeneration Protocol for the Native Grass Polypogon australis Brong

Polypogon australis (Brong.), a native Chilean grass (Poaceae), is a facultative metallophyte capable of colonizing copper mine tailing dams and adapting to saline and acidic substrates. These traits make it a promising candidate for phytoremediation of metal-contaminated soils. However, the lack of in vitro propagation, callus induction, and somatic embryogenesis protocols limits its use in large-scale applications and genetic improvement. This work aims to establish a reproducible in vitro regeneration system for P. australis . Mesocotyl explants were cultured on Murashige and Skoog (MS) medium supplemented with 2.5 mg L ⁻¹ Dicamba. Callus induction was achieved in 38.9% of explants, and 45.5% of embryogenic calli regenerated into plantlets producing leaves and radicles without requiring exogenous organogenesis-inducing hormones. The regenerated plants continued to develop further on MS + sucrose medium, confirming the totipotent capacity of mesocotyl-derived calli. The developed protocol provides a foundation for large-scale propagation and genetic transformation of P. australis . By overcoming propagation bottlenecks, this methodology strengthens the potential of this native metallophyte as a model for phytoremediation and future CRISPR-based biotechnological approaches to enhance copper tolerance and accumulation.