Embryogenesis and synthetic seed production in Mondia whitei

Abstract

An effective plant regeneration system via somatic embryogenesis and synthetic seeds was developed for Mondia whitei, an endangered medicinal plant. Friable embryogenic callus was induced by culturing leaf explants on solid Murashige and Skoog (MS) medium containing various concentrations and combinations of sucrose and plant growth regulators. The highest frequency of somatic embryogenesis (100 %) and production of all developmental stages of somatic embryos were obtained on MS medium with 40 g l−1 sucrose, 8 g l−1 agar, 20 μM 2,4-dichlorophenoxyacetic acid (2,4-D) and 1 μM thidiazuron. This was followed by establishment in MS medium with 20 g l−1 sucrose, 8 g l−1 agar, 0.5 μM meta-topolin riboside (mTR) and 0.25 μM indole-3-acetic acid (IAA). All the embryos germinated and produced healthy plantlets on the same medium. Somatic embryos at the heart, torpedo and cotyledonary-stages were collected from media (EDM) containing MS medium plus 20 g l−1 sucrose, 8 g l−1 agar, 0.5 μM mTR and 0.25 μM IAA. The embryos were encapsulated with liquid MS medium plus different concentrations of sodium alginate (SA) and calcium chloride (CaCl2·2H2O) with a 10 min exposure. A combination of 3 % SA and 100 mM CaCl2·2H2O provided higher survival (95.7 %) and germination (73 %) frequencies of synthetic seeds. Germination frequency of synthetic seeds was 51.6 % after 50 days of storage at 4 °C. Somatic embryos and synthetic seed-developed plantlets were successfully acclimatized in the greenhouse with 90 % survival ex vitro. Application of the protocol provides a relatively simple and rapid system for conservation of natural populations for germplasm conservation. Analysis of bioactive compounds and genetic transformation studies can also be performed.