Role of Thidiazuron in Modulation of Shoot Multiplication Rate in Micropropagation of Rauvolfia Species

Abstract

Thidiazuron (TDZ) is a light yellow crystalline phenylurea compound with tremendous morphogenic potential on shoot regeneration ranging from small grasses to large tree species. This morphoregulatory potential of TDZ has led to well-established micropropagation systems in various woody plant species where other cytokinins are least effective. Short exposure of TDZ is sufficient to stimulate the best regeneration system, while the prolonged or continuous exposure has various negative effects on growing cultures. The present communication reviewes the morphogenic response of TDZ in Rauvolfia species. A differential growth response was observed on explants when inoculated on Murashige and Skoog (MS), 1962 medium supplemented with different concentrations of TDZ. Nodal explants of R. tetraphylla cultured on MS + 5 μM TDZ gave optimal (90%) regeneration response for maximum (9.2 ± 1.20) shoot production, while in the case of R. serpentina, MS + 0.8 μM TDZ proved to be best for producing highest mean shoot number (5.7 ± 0.28) with 77% regeneration rate after 4 weeks of culture. However, for shoot tip explants of R. hookeri, 0.45 μM TDZ on ½ MS proved best and gave 70% regeneration, with a maximum shoot number of 1.50 ± 0.22 and a mean shoot length of 1.18 ± 0.04 cm after 45 days of culture. Responsive nodal explants of R. tetraphylla and R. serpentina when cultured continuously on the same TDZ-supplemented media lead to adverse effects like shoot distortion, fasciation, or hyperhydricity in growing shoots and resulted in stunted growth. These negative effects of prolonged TDZ exposure were apprehended when the cultures were transferred to MS basal medium devoid of TDZ. The transference of cultures to secondary medium not only showed the positive effect on growing cultures but also increased shoot proliferation and shoot multiplication rates in both the species. After 4 weeks of transfer to secondary medium, shoot number increased up to 23.1 ± 0.4 and 18.5 ± 1.25 per explant in R. tetraphylla and R. serpentina, respectively. Regenerated shootlets of ≥4 cm were excised and transferred to various rooting medium supplemented with different concentrations of auxins like IAA, IBA, and NAA. IBA in all the three species proved best for in vitro rooting with maximum root mean number of 6.9 ± 0.34 and mean root length of 5.2 ± 0.4 cm at 1.0 μM IBA in R. serpentina after 4 weeks of incubation. Rooted plantlets were acclimatized in culture room and finally transferred to garden soil with 90% survival rate without any genetic or morphogenic abnormality.