Abstract
Induction of somatic embryogenesis and complete plantlet regeneration from callus culture of Hibiscus sabdariffa L. var. HS4288 has been made. Leaf and root explants were cultured on Murashige and Skoog (MS) and Driver–Kuniyuki Walnut (DKW) basal media supplemented with different concentrations of synthetic auxins and cytokinins. Root explants on DKW medium supplemented with 2.26μM 2, 4-Dichlorophenoxyacetic acid (2, 4-D) and 4.65μM kinetin (KIN) induced highest percentage (70%) of embryogenic calli. Average number of globular embryos per root derived callus produced within 6 weeks of culture initiation on MS media with different plant growth regulators (PGRs) ranged from 2.27±0.12 to 8.80±0.17 and that of cotyledonary embryos ranged from 0.00 to 2.53±0.20. On DKW medium comparatively more globular embryos (2.70±0.15 to 14.53±0.23) and cotyledonary embryos (0.00 to 8.90±0.17) were produced than that of MS medium. Regeneration of complete plantlets was highest (76.67%) when embryogenic calli with mature somatic embryos were grown on DKW medium containing 2.32μM KIN and 2.22μM 6-Benzyladenine (BA). Plants were primarily hardened in humidity, temperature and light controlled chamber and finally in a greenhouse showed 70% survival ability. Different stages of somatic embryogenesis process in the root derived embryogenic calli were elaborated in detail by morphological, histological and SEM study. The data were statistically analyzed by Duncan Multiple range test (p ≤ 0.05) and Principal component analysis (PCA). Flow cytometry and Inter-simple sequence repeats (ISSR) marker analysis confirmed that there was no genetic variation within the regenerated plants.
Highlights
Hibiscus sabdariffa L., an annual plant of family Malvaceae is cultivated in India and many other tropical and subtropical countries
To delineate the cumulative distribution of two different types of explant for the induction of embryogenic callus on Murashige and Skoog (MS) and Driver–Kuniyuki Walnut (DKW) culture medium a concerted effort has been made by the use of principal component analysis (PCA)
In an earlier publication of somatic embryogenesis in two varieties of H. sabdariffa [13], it was reported that the root was not suitable explant for embryogenesis study
Summary
Hibiscus sabdariffa L., an annual plant of family Malvaceae is cultivated in India and many other tropical and subtropical countries. Due to the cleistogamous flower of this plant, conventional hybridization is restricted As it is a tetraploid plant (2n = 72) [6], segregation in the generation and purification of the population require a prolonged time in conventional approach of genetic improvement. Sylvere Sie et al [13] were the first to report somatic embryogenesis from hypocotyl and cotyledon-derived calli; the root derived calli showed poor response to the induction of somatic embryos [13]. In this background, our objective was to test the possibility of induction of somatic embryogenesis from root and leaf derived calli of H. sabdariffa L. var. We studied the different stages of somatic embryo development, embryo maturation and their differentiation into complete plants as well as their acclimatization under controlled environment in the lab and in the field
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