Study On Leaf Culture And Octoploids Induction Of Cultivars Phalaenopsis

Phalaenopsis is an important economical flowering plant. In recent years, with promotion of species breeding selection and R&D of the industrialized production , the number of new species, production scale and market share have been increasing. However, the industrial development of Phalaenopsis is obstacled by long production cycle, low efficiency of tissue culture propagation and less available resource variety. Therefore, there is great urgent to study on tissue culture propagation methods and germplasm innovation. On the one hand,through the experiment on 9 kinds of Phalaenopsis genotype materials, this study is discussing the impacts on adventitious shoots induction and plantlet regeneration of the influence factors of tissue culture, including position of explant, basic medium, proportion of cytokinin and auxins, the dark treatment, leaf size, organic additive and genotypes. On the other hand, the study discusses two methods of octoploid Phalaenopsis adventitious shoot induction by colchicine treated and finds out the appropriate induction concentration and processing time. These results could provide useful reference for new species breeding and large-scale production of Phalaenopsis. The research results are as follows:1 Six varieties were cultured for adventitious shoot induction in NDM and 1/2MS, the results show that induction rate of different varieties have distinct difference in the same medium, the induction rate of 0377-2 is 25.0% , but Dtps.Queen Beer'Red Sky',P.Hehuang,P.Taida Salu is 0% in 1/2MS; adventitious shoot induction rate of the same varieties also have distinct in different medium. 1/2MS is more suitable medium for adventitious shoot induction than NDM.2 The leaf and root of 0377-2 were cultured in the mix medium with 1/2MS + 3mg/L TDZ,the leaf induction rate is 34.9% with characters of short inducing time of 2 mouths, the root induction rate is 13.3% with long inducing time of 3-4 mouths; the induction time of the leaf is shorter than the root; and the root induction is more difficult than the leaf induction. In addition, adoption of 1/2MS+3mg/LTDZ and supplementation of 200ml coconut water were found to help to facilitate callus induction.3 The leaves were respectively cultured on 1/2MS medium supplemented with TDZ,6-BA, KT-30,Ad,NAA,IBA and the composition. Effects showed that the optimum effect comes from the treatment of TDZ, the next is 6-BA, the effect of KT-30 and Ad is worst among cytokins ; the effect of TDZ or 6-BA treatment alone was better than combined of NAA; The adventitious shoots induction rate reached 100%,70.3%,31.7% for the treatment of TDZ, 6-BA, NAA, respectively; while the adventitious shoot induction rate reached 96.7% and 33.3% for the combined of TDZ+NAA and 6-BA+NAA, respectively. By the treatment of TDZ 3mg/L, P.Tailin'Red Angle'induction rate reaches 100% and the average number of adventitious shoots reaches 18.2. However, TDZ significantly inhibited the growing of adventitious shoots and the rooting of plantlet, IBA with 1mg/L—3mg/L can accelerate the growing and the rooting of plantlet.4 The results also show that less than 15 days of dark treatment and leaf shorter than 1cm as explants were suitable for adventitious buds induction from the leaves.5 We adopted two kinds of colchicine treated to induce polyploid from the leaves. The first Colchicine-Treated: After adventitious buds induction from the leaves of one month on the medium of 1/2MS + 1mg/L TDZ, the leaves with small new formed buds are soaked in different concentrations solution to induce polyploid. The second Colchicine-Treated: After adventitious buds induction from the leaves for some days on 1/2MS+1mg/L TDZ+ colchicine, then the leaves with small formed buds were transferred to the medium of 1/2MS+1mg/L TDZ to obtain the reduplical regeneration plant. The results show that the first Colchicine-Treated is harmful for explants itself, but it is more effective for polyploid induction. The induced frequencies of octoploid is up to 33.3% when the explants was cultured for 30 days in the treatment 0.01% colchicine with the first Colchicine-Treated, but the induced frequencies of octoploid is up to 6.32% when the explants was cultured for 5 days in the treatment 0.1% colchicine with the first Colchicine-Treated.P.Tailin'Red Angle'was induced to octoploid plant by the treatment of colchicine. The basic characteristics of octoploid includes 2n=8x=152 of chromosomes, slower growth, thicker leaves, shorter and stouter roots, bigger and thinner stomata density.