| Paphiopedilum, due to excessive exploitation, habitat destruction; and difficult propagation, has become one of the most endangered plant species in the world. P. Maudiae Type are a group of hybrids~with great value in the market, their cultivation and management are relatively more easier than native species or other hybrids. At present,the main propagation method of P. Maudiae Type is aseptic seed germination, which can not keep the excellent characteristics of parents, the clonal reproduction is very important method in commercial production. However, the tissue culture of the Paphiopedilum is very difficult, and there is no report on the scale seedling production by tissue culture. Therefore, this thesis focuses on the tissue culture of P. Maudiae Type, in order to provide the basis for the mass micropropagation of Paphiopedilum and strengthen the protection of native species of paphiopedilum.In this paper, the tissue culture of different explants from asymbiotic germination of P. Maudiae Type and transplanting of plantlets by tissue culture were studied. The main results were indicated as follows:1. Thin layer of protocorm culture: longitudinal sections of protocorms were more suitable explant for adventitious bud induction than transverse section. Longitudinal sections cultured in the culture medium 1/2 MS+1.0 mg/L BA+0.5 mg/L NAA could form many buds. The callus was not induced by protocorm cultured. In the subculture,1/2 MS +5 mg/L KT was more suitable for PLB formation and proliferation. Under long dark treatment, the induction rate of PLB was very low. In liquid suspension culture, protocorm only changed in morphology, and did not proliferated.2. Cluster bud induction and proliferation from seedling explants: Cluster bud induction and proliferation was the main method of micropropagation of P. Maudiae Type, 1/2 MS+0.5 mg/L TDZ was better to promote the proliferation of seedlings at the beginning of culture. In the subsequent subculture 1/2 MS+5 mg/L BA could promote seedling growth. Combined with the culture of multiple generations of cluster buds, 1/2 MS+5 mg/L BA +0.5 mg/L NAA was most suitable medium for cluster bud proliferation. The leaves was not suitable to be used as explants for tissue culture because of their browing and dying.3. Seedling growth in vitro: After three months of culture, the plant height, leaf number and root number were measured: 20 g/L sucrose concentration treatment can significantly increase the seedling height and root number. 1 g/L H1 +1 g/L H2 was most appropriate basic medium for seeding growth, the number of expansion leaf was more and the plant height was higher than other treatment.4. Seedlings transplanting: coconut shell used individually or in combination with other medium was not suitable to the growth of plantlets transplanted. The bark used alone achieved the highest survival rate 92.5%, which could be used as the transplanting medium of tissue culture seedling. However, water~retaining capacity of the bark was poor and should be watered more often. Meanwhile, and its density was small, was not conducive to the management of large seedlings. Zhijing stone∶bark( V/V=2∶1) was suitable mixed~matrix for transplanting and management of large seedlings. |
