Studies On In Vitro Flowering Of Dendrobium Nobile Lindl

Wild Dendrobium nobile Iindl take several years to flower. In vitro flowering by plantlets produced through tissue culture can decrease the juvenile phases from a vegetative to a reproductive stage. Studing molecular mechanisms of D. nobile Lindl floral induction, floral meristem formation, and floral organ development are important not only in theory but also in application.The objective of this research is to study the effects of PP333, ABA, TDZ, BA, nitrogen concentrations and sucrose concentrations on flowering in vitro of D. nobile Lindl. The results of our research show that BA had an indistinctive effect on in vitro flowering of D. nobile lindl. The highest frequency of floral bud induction was about 34.3% if induced by TDZ only. 1/2MS medium with less nitrogen sources (1/10N) and more phosphor concentration (5P) and supplemented with 40g/L sucrose could make the frequency of floral buds induction increased greatly reaching 76.3% under the induction of TDZ. Effects of PP333 or ABA pretreatment on in vitro flowering of D. nobile plantlets induced by TDZ were not obvious. The number of floral buds was increased if pretreated by "PP333+ABA". The optimal way for inducing floral buds is to preculture the plantets on 1/2MS medium supplemented with 0.5mg/L ABA and 0.5mg/L PP333 for 35 days, then transferred them onto 1/2MS medium supplemented with 1.0mg/L PP333 and 0.1mg/L TDZ. The frequency of floral buds induction increased sharply reaching 62.2% during a period of 150 days. All the mediums were added to 30g/L sucrose and 9g/Lagar. The pH was adjusted to 5.6-5.8 with IN NaOH. In addition, the frequency of floral bud induction was also affected by root excision. Most of the plantlets able to form floral buds were not the original ones but those grew up from new stems. On the top of them, a single flower or a inflorescence including 2~3 small flowers could be observed. MS medium supplemented with 0.5mg/L NAA and different concentrations of PP333 could make D. nobile plantlets thrived. Floral buds were also formed during the process of thriving treatment. If thriving the plantlets before precultured, the frequency and the sum of normal flowers were sharply raised atthe same conditions of pretreatment and induction.After having screened the optimal medium for inducing flower buds initiation on D. nobile plantlets, we observed the process of early stage of floral bud differentiation. The whole process was divided into 8 stages. After 20 days of culture, the floral bud was still undifferentiated and it would differentiate in 30 days. Floral anlage was formed after 40 days of culture. After that, sepal anlage, petal anlage and habenaria reniformis appeared gradually after 50-80 days of culture.The components of the proteins were analyzed by SDS-PAGE. When plantlets were cultured in comparison mediums, two special proteins about 36KD and 29KD appeared in 30 days, and a special protein about 12KD appeared in 75 days. In addition, three special proteins about 54KD, 13KD and 12KD appeared when plantlets were cultured in the optimal medium for 90 days, and another three special proteins about 82KD, 42KD and 28KD could be observed after cultured in the same medium for 105 days.