Exogenously added polyamines recover browning tissues into normal callus cultures and improve plant regeneration in pine

Polyamines are known to influence a variety of growth and developmental processes in higher plants. Tissue browning seriously reduces in vitro plant regeneration in pine species by decreasing the levels of antioxidant enzymes and polyamines in tissue cultures. In the present investigation, the effect of exogenously added polyamines on recovering browning tissues into normal callus cultures and on improving plant regeneration was examined in Virginia pine (Pinus virginiana Mill.). Among the polyamines administered, 1.5 mM putrescine (Put), 1.5 mM spermidine (Spd), or 1.5 mM spermine (Spm) alone resulted in a 19.55%, 18.92%, and 1.45%, respectively, recovering of browning tissues into normal callus cultures in 5 weeks. A combination of Put with Spd or Spm did not result in an increase of recovering rate, compared to the Put or Spd alone. Exogenously added 1.5 mM Put, 1.5 mM Spd, 1.5 mM Put + 1.5 mM Spd, or 1.5 mM Put + 1.5 mM Spm recovers browning tissues into normal callus cultures by increasing the activity of antioxidant enzymes ascorbate peroxidase (APOX), glutathione reductase (GR) and superoxide dismutase (SOD) and by decreasing lipid peroxidation. Exogenously added 1.5 mM Put, 1.5 mM Spd, 1.5 mM Put + 1.5 mM Spd, or 1.5 mM Put + 1.5 mM Spm significantly improve the growth rate of callus cultures, shoot formation, and rooting adventitious shoots. These results demonstrated that exogenously added polyamines recover browning tissues into normal callus cultures by decreasing oxidative damage and improving plant regeneration by acting as plant growth substances.