14C-2,4-DICHLOROPHENOXYACETIC ACID UPTAKE AND FORMATION OF EMBRYOGENIC CALLI IN COCONUT PLUMULAR EXPLANTS CULTURED ON ACTIVATED CHARCOAL-FREE MEDIA

Abstract

Research for the development of protocols for the micropropagation of coconut plants (Cocos nucifera L.) has encountered several difficulties, such as explant browning. Although browning could be overcome by adding activated charcoal, this additive can also bind components of the culture medium required for the tissue morphogenic responses; this adsorption can be up to 99 % of the growth regulators added to the medium. In this work we developed a protocol without the use of activated charcoal which is able to produce embryogenic calli from coconut palm plumules as explants. This protocol also allowed us to study the kinetic absorption of 2,4-dichlorophenoxyacetic acid (2,4-D) by the explant, otherwise impossible in medium with activated charcoal. After evaluating different concentrations of 2,4-D in culture media without charcoal or polyvinylpolypirrolydone (PVPP) instead, we found that about 43 % of coconut explants cultured in media containing 1 µM 2,4-D without
activated charcoal (with or without PVPP) formed embryogenic calli after four months. Yields were reproducible and similar to those obtained with medium containing activated charcoal and responses occurred more rapidly. Morphological observations of calli obtained in activated charcoal-free media were similar to those obtained in activated charcoal-containing medium. However there were differences in the histological observations, mainly in the shape of the nuclei of the proembryos of both types of callus. Uptake of ¹⁴C-2,4-D was studied in plumule explants cultured in activated charcoal-free medium. After 120 d of culture about 25 % of the original radioactivity originally in the medium was extracted from explants and their ¹⁴C-2,4-D concentration peaked within the first 20 d, before appearance of any morphogenic response.