Efficient culture protocol for plant regeneration from cotyledonary petiole explants of Jatropha curcas L.

A high-frequency and reproducible protocol for induction of adventitious shoot buds and plant regeneration from cotyledonary petiole explants of Jatropha curcas L. has been developed. The cotyledonary petiole explants of J. curcas cultured directly on medium supplemented with thidiazuron (TDZ) induce regeneration of poor quality shoot buds that have a low regeneration frequency. However, treating the explants with high concentrations (10-60 mg/L) of TDZ solution for certain time periods (5-80 min) significantly increased the regeneration frequency and improved the quality of the regenerated shoot buds. The best shoot buds induction (88.42%) and number of shoot buds (12.67) per explant were observed when in vitro explants were treated with 20 mg/L TDZ solution for 20 min before being transferred on hormone-free medium after 30 days. Regeneration was also influenced by the orientation (horizontal or vertical) of the explants on the medium, and by the origin of the cotyledonary petioles (in vitro or in vivo) used for the preparation of explants. We performed subsequent experiments for elongation and rooting of the regenerated shoot buds. Addition of L-arginine to the medium was conducive to the elongation of the shoot buds. A concentration of 7.5 mg/L L-arginine yielded the best results. The elongated shoots could initiate roots to become intact plantlets in half-strength Murashige and Skoog medium containing 0.1 mg/L indole-3-butyric acid. After acclimatization, these plantlets could be transplanted to the soil and the growth was normal. Therefore, application of the methods described here helped to increase plant regeneration efficiency.