In vitro regeneration of four Hypoxis species and transformation of Camelina sativa and Crambe abyssinica

Abstract: The advance of science and biotechnology has made it possible to prevent the complete loss of plant species by providing propagation methods that are fast and can result in the mass production of exact copies of the targeted plant species. Genetic transformation has become a useful instrument in the introduction of new traits to plants such as increased yield and disease resistance. In this study potential in vitro regeneration protocols for four Hypoxis species were successfully developed. Efficient indirect regeneration (100% with 8 shoots per explant) of Hypoxis filiformis was obtained when corm explants were directly cultured in MS supplemented with 1 mg/l NAA. However, the highest mean number of shoots per explant (17) was obtained in basal MS supplemented with 3 mg/l kinetin. Efficient direct regeneration of H. acuminata (100% with 2 shoots per explant) was achieved in corm explants cultured with a piece of shoot attached in MS supplemented with 3 mg/l kinetin. Up to 30% and 21% seed germination was obtained in H. argentea and H. filiformis respectively, when seed coats were crushed before culturing in half-strength MS without PGRs. In addition, the use of activated charcoal and 0.1% mercuric chloride, respectively, helped in controlling the build-up of phenolic exudates in the medium and infection of explants and shoots by endogenous micro-organisms. Agrobacterium-mediated transformation of Camelina sativa and Crambe abyssinica flowers by vacuum infiltration with three gene constructs: AtHb2. BvHb2, and vhb in camelina; and the FWS 3-1 vector harbouring three genes: ScFAR, ScWS, and DsRed in crambe failed to produce transgenic seeds.