Optimization of a Mercury-free Surface Sterilization Protocol for Dried Sorghum Grains (Sorghum bicolor [L.] Moench): Balancing Contamination Control and Explant Viability

Introduction: Aseptic culture establishment is essential for plant tissue culture and genetic transformation. In sorghum ( Sorghum bicolor [L.] Moench), seed-associated microorganisms frequently compromise culture initiation and reduce regeneration efficiency. Sodium hypochlorite (NaOCl) is widely employed as a disinfectant, but optimal application conditions for dried sorghum grains remain poorly characterized. Methods: This study investigated the interaction between NaOCl concentration and exposure duration on contamination and explant survival. Sixteen treatment combinations of four NaOCl concentrations (20%, 30%, 40%, and 50% commercial Clorox®) and four exposure durations (10, 15, 20, and 25 min) were tested in a factorial randomized complete block design with fifteen explants per treatment (240 total). Sterilized grains were cultured on hormone-free Murashige and Skoog medium and evaluated over two weeks using two-way ANOVA and binary logistic regression. Results: Both concentration and exposure time had a statistically significant effect on contamination and viability, with a statistically significant interaction between them. 20% NaOCl for 25 min was the optimal condition with zero contamination (0.067 ± 0.064) and 100% viability (1.000 ± 0.000). High concentrations yielded greater inhibitory effects but decreased viability. The optimum conditions as estimated from logistic regression were near 21.6% NaOCl and 24.3 minutes, near the optimal empirical conditions. Conclusion: The results indicate that the use of concentrated disinfectants or mercury-based compounds is not necessary for good sorghum grain disinfection. The use of moderate concentrations of sodium hypochlorite for relatively long periods offers acceptable antimicrobial activity and low phytotoxicity. The protocol outlined is viable and eco-friendly for sorghum tissue culture and could be useful for improving the future regeneration and transformation of this economically important crop.