Modification of In Vitro Culture Method of Paphiopedilum glaucophyllum for Callus Induction

Modification of In Vitro Culture Method of Paphiopedilum glaucophyllum for Callus Induction

The in vitro vegetative propagation technique of the Paphiopedilum glaucophyllum (callus propagation) still haveproblems to date. The aim of this study is to determine the effect of the media and plant growth regulator (PGR) on seed germination andcallus induction with shoot tip as an explant and use the planlet from elongation treatment result. The seed germination uses modifiedKnudson C (KC medium) with the addition of 3 concentrations of NAA (0; 5; 10 mg/L). The second study is the elongation treatmentof the explants uses four modified Murashige and Skoog (MS) medium (1/2 P, ½ P5, MP and MP5). And for the callus induction studyconsisted of two methods, callus induction with direct planting to the treatment medium by using SH medium with addition of2,4-Dichlorophenoxyacetic acid (2,4-D) 1 mg/L and Thidiazuron (TDZ) (0, 0.5, 1, 1.5 dan 2 mg/L). The second method used theexplants resulted from elongation treatment. It used half-strength Wattanawikkit medium with addition of TDZ (0; 0.5; 1; 1.5; 2 mg/L)and 2,4-D (0; 1; 5; 10 mg/L). All callus then subcultured in the half-strength MS (½ MS) medium containing 2,4-D 1 mg/L and TDZ (0;1; 1.5 mg/L). All the experiments used a completely randomized design with 3 repetitions. The results showed that the seeds germinated2 months after planting (MAP) for all media. The optimal media for explants elongation of P. glaucophyllum was MP5 media. Meanwhile, for the first callus induction experiments, explants had more callus in basic SH media with 2.4-D 1 mg/L and TDZ 0.5-1.5mg/L that grew in the dark. The second experiment, the optimal medium for callus induction was half MS with Thidiazuron 1.5 mg/Land 2.4-D 1 mg/L (T1.5D1). The the suitable medium for callus development is ½ MS) medium with 2,4-D 1 mg/L.