Somatic embryogenesis in pearl millet (Pennisetum glaucum): Strategies to reduce genotype limitation and to maintain long-term totipotency

Three genotypes of Pearl millet were screened in vitro for induction of emb ryogenic callus, somatic embryogenesis and regeneration. Shoot apices excis ed from in vitro germinated seedlings or immature embryos isolated from gre en house established plants were used as primary explants. The frequency of embryogenic callus initiation was significantly higher in shoot apices in comparison with immature zygotic embryos. Moreover, differences between gen otypes were minimal when using shoot apices. Friable embryogenic calli (typ e II) developed on the initial nodular calli after 1 to 3 months of culture . The frequency of type II callus is related to the composition of the main tenance medium and they were more often found in ageing cultures. The trans fer of embryogenic calli onto auxin-free medium was sufficient for inducing somatic embryo development in short-term culture (3 months) while a progre ssive loss in regeneration potential was observed with increasing time of s ubcultures. Maturation of embryogenic calli on medium supplemented with act ivated charcoal, followed by germination of somatic embryos on medium suppl emented with gibberellic acid, restored regeneration in long-term cultures.