Tissue Culture And Establishing Agrobacterium-mediated Transformation System In Cymbidium Goeringii Rchb.f.

The widely cultivated orchids in China belong to Cymbidium, and are all perennial and evergreen herbaceous plants. There are approximate 40 species in Cymbidium, which are mainly distributed in the tropic and subtropic areas of Asia, and some in Oceania and Africa. China has 20 species and mutations. Orchid is one of China's traditional and famous precious flowers, So, it is great potential for study on orchid.This experiment take Cymbidium goeringii Rchb.f. , as explants, studied systematically many factors' effect on proliferation, growth, radication and comeback after long time in vitro culture on shoot and PLB. At the same time, an efficient Agrobacterium-mediated transformation system for Cymbidium goeringii was established. The primary results were summarized as follows:(1) Healthy green PLB were obtained from shoots and buds on the MS medium supplemented with 5. 0 mg/L NAA. We explored deeply on the optimal proliferation medium, and ascertained the suitable concentration of BA (0. 1 mg/L). We also compared the effects of seven organic additives on the increase of fresh weight and proliferation of PLB, indicating that both Yeast additives and peptone were the suitable additives, and 1. 0 g/L peptone showed the most effective.(2) During the culture of shoots, through adjusting the concentrations of NAA, not only the buds could be induced to form multiple shoots, but also the proliferating rate of the buds was improved. Simultaneously it was found that the height of shoots was critical for rooting. Sucrose at suitable concentration (20 g/L) was also necessary for the normal growth of C. goeringii..(3) In the process of proliferation of PLBs and weakly-growing shoots, F/P rate (growth rate of flesh weight / proliferation rate of PLBs) was a key parameter that could be used to distinguish whether the growth of PLBs was normal or not. When F/P rate was not less than 0.6, PLBs showed well growing and developed, otherwise weakly growing and showing greenwish to yellowish. And for shoots, the suitable F/P rate was around 1. 12.(4) A system of Agrobacterium-mediated transformation for C. goeringii. was established. GUS histochemical assay and PCR analysis showed that gus gene was integrated into C. goeringii successfully.