Establishment Of Phalaenopsis In Vitro Propagation And Its Polyploidy Induction

Phalaenopsis amabilis is a kind of luxurious flower with important admire value and economic value. It has been widely cultivated in the world, and has a vast marketplace prospect. For the industrialization of Phalaenopsis in our country, it is very urgent to establish and optimize its rapid propagation technology.In this paper, we studied three Phalaenopsis species in vitro tissue culture system. In addition, in order to establish the induction and identification technology of Phalaenopsis polyploidy, we take the mini-series Phalaenopsis as the material and take to induce its ployploidy with the colchicine mutagen. The main results are as follows:1. The rapid propagation of Phalaenopsis cluster shootsThe pedicels with axillary buds were chosen as the explants to induce the clustered shoots. The results showed that the surface steritization method of treatment in 70% alcohol solution for 30s and then in the mixture of 0.1% HgCl₂ and several drops of Tween-20 for 12min gave a survival of 91.67%. On medium of 1/2MS+ BA3.0mg/L +NAA0.2mg/L or l/2MS+BA6.0mg/L, the induction rate of axillary buds reached 100%. When the axillary buds were cut down and trasffered on the medium of l/2MS+BA3.0mg/L, the clustered shoots were induced in a month. The orthogonal test proved that the medium of l/2MS+BA5.0～10.0mg/L+NAA0.5～1.0mg/L+glucose or sugar was the optimal medium for clustered shoots propagation.2. The rapid propagation of protocorm-like body from Phalaenopsis leavesIn this research, the effects of the dark culture, the leaf explants ages and the culture media on protocorm-like body induction were studied. The results showed that when the leaves were preculture in dark condition for 14 days and then shifted to light culture, it was benefit for protocorm-like body induction. Young leaves were the better explants for the induction. For older leaves, they should be cut into the blocks when chosen as the explants and the leaf base was the best position to induce protocorm-like body. The medium of modified MS+BA10.0mg/L+NAA1.0mg/L was suitable for protocorm-like body induction, and the medium of 1/2MS+BA3.0mg/L+NAA0.5mg/L was optimal for the proliferation.3. Rooting of tube seedlingsDuring the adventitious root formation, the 1/2MS medium supplemented with a little NAA gave a good result. In this experiment, the optimal medium for rooting was l/2MS+NAA1.0mg/L.4. Transplanting and management of tube seedlingsThe seedlings with 3～4 leaves and 2～3 roots were transplanted. The results showed that acclimatization for 4～5 days and treatment with Carbendazin for 10～20min or potassium permanganate for 20 min increased the seedlings survival. The seedlings in substrate with sphagnum, peat and perlite gained a survival rate of 100%. On the other hand, the seedlings survival rate in water culture declined to 64.6%, but the survivaled seedlings had a better growth potential.5. Polyploidy inductionDifferent colchicine concentrations and treatment durations had different effects on the survival and proliferation rate of young shoots. Higher colchicine concentrations resulted in a lower survival rate and proliferation rate. When treated for 48h with 0.2% colchicine, the survival rate was 60%.The chromosome mumber were counted by the improved card-stained magenta compression method, and it showed that the treatment for 24h with 0.05% colchicine gave a induction rate of polyploidy of 20%. In this study, four tetraploid plants, two octoploid plants were obtained. Tetraploid plants(2 n = 4x = 76) were characterized by higher growth potential and thicker leaves with darker color. In addition, the stomata lengths on the lower leaf surface of tetraploid plants were notably longer than those of the diploid, while the stomata density was obviously decreased.