In vitro plantlet regeneration in Annona squamosa through direct shoot bud differentiation on hypocotyl segments

Abstract

A highly reproducible in vitro regeneration system in Annona squamosa using hypocotyl segments has been developed. Multiple shoots were induced in vitro from hypocotyl explants through adventitious shoot bud regeneration. The ability of hypocotyl segments to produce shoot buds varied depending upon their position on the seedlings. Hypocotyl segments obtained from closer to the root were highly responsive and produced a maximum of 23 shoot buds per explant on MS medium supplemented with 5.0 mg l −1 BAP. TDZ at all the concentrations tested led to the formation of a large amount of callus. Incorporation of auxins NAA, IAA, IBA or 2,4-D during shoot bud induction did not show any improvement over that obtained with BAP and promoted callusing to a different degree. Shoot buds produced on BAP medium, however, remained stunted. Reduction in concentration of BAP from 5.0 to 2.0 mg l −1 after two subcultures improved the shoot growth and elongation. Callus free rooting in 70% of shoots was obtained when they were treated with pre-autoclaved IBA (200 mg l −1) for 45 min and directly implanted on autoclaved Soilrite™ contained in culture bottles and irrigated with one-fourth MS salt solution. This allowed rooting and partial hardening simultaneously. Complete hardening was carried out in a greenhouse with more than 70% success.