Improved protocol for efficient regeneration of coconut (Cocos nucifera L.) anther derived embryos

The occurrence of severe shoot necrosis and other constraints, such as low frequency of embryo induction and poor regeneration of plants, restricts the use of coconut androgenesis in practice. Fine-tuning of the protocol by addressing the above constraints was carried out with the intention of scaling up haploid plant production. Out of the carbon types, sucrose and maltose when added in concentrations of  90.0 gL^-1 and 120.0 gL^-1 showed significantly higher (p < 0.05) embryo (44.0 % and 36.0 % respectively) production. Out of the concentrations used in the study, 20.0 μM 6-benzylaminopurine BAP showed significantly (p = 0.001) higher shoot generation (47.6 %) as well as significantly (p = 0.006) longer shoot production (31.7 %) within the study period. The effect of CaCl₂ on the reduction of shoot necrosis was also tested. CaCl₂ showed a significant (p = 0.001) effect on reducing shoot necrosis. The lowest occurrence of shoot necrosis (25.0 %) was observed in 4.0 mM CaCl₂ treatment. Continuous sub-culturing of shoots with initial signs of shoot necrosis, to elevated CaCl₂ levels until the rooting stage, facilitated the recovery. The transfer of shoots frequently in to a fresh medium was not beneficial for the suppression of necrosis. Shoots maintained in the medium enriched with 4.0 mM CaCl₂ were transferred for acclimatisation, and this is the first report of transferring haploid coconut plants to acclimatisation conditions. The rooted shoots produced through the optimised protocol were acclimatised successfully. The prevention of shoot loss due to shoot necrosis will be beneficial for further refinement of the coconut anther culture protocol.