Studies On Tissue Culture And Genetic Transformation Of Acacia Crassicarpa

In this paper, we studied tissue culture of Acacia crassicarpa systematically. The system of tissue culture and agrobacterium-mediated transformation of Acacia crassicarpa was established and some GUS transgene plantlets of Acacia crassicarpa had been obtained successfully. The results were included in the following:1, Amend 1/2MS medium without any hormones was suitable to sterilite seeds of Acacia crassicarpa and amend MS basic medium tissue culture.2, 0.1% HgCl₂ 8 min + 10% NaClO 20min was suitable to sterilited the seeds, the contamination rate was 0 and the rate of seeds aprout 89%.Seeds sterilition was uncompleted by using NaCIO singly.3, The pathway of adventitious bud differentiation may become the main pathway on tissue culture of Acacia crassicarpa. The differentiation rate of callus produced by the pathway of differentiation was low, which was disadvantage to tissue culture of Acacia crassicarpa.4, 2, 4-D and KT not fit for induction of callus, buds and plantlets, but 6-BA can promoted inducting callus and plantlets. The best medium of buds induction was amend MS+6-BA_1.0-2.0mg/L +IBA_0.2mg/L. The highest buds multiplication times was 4.03.5, Adding 10mg/L CM in the media can promote buds induction.6, Cotyledon age of seedling 30d was the best explant age for the tissue culture of Acacia crassicarpa's seeds.7, 20 days/generation was favourable for the multiplication of buds and plantlets.8, Ability of buds multiplication was stronger and longer. The buds can subculture more than 8 times continually.9, In the culture of root inducetion and sound seedling, the optimal medium was MS medium supplemented with 0.5-1.5mg/L Met.10, Agrobacterium of A208SE, At05, At06 couldn't grow and explant differentiation rate could keep higher when Tb concentration was 250mg/L.11, The sensitive concentration of Km was 50-75mg/L on the leaf-fragment of Acacia crassicarpa. The non-transgenic plantlet couldn't induce roots on the medium contained Km.12, The leaf-fragment of Acacia crassicarpa needn't pre-culture, GUS gene instantaneous expressed perfectly. The rate of instantaneous expression was 10%.13, Leaf-fragment of Acacia crassicarpa were dipped in agrobacterium liquid that concentration was OD≈0.3-0.5 for 10-15 min and co-cultivated in three days, the GUS gene expressed optimal and the expressed rate was 26.7%.14, GUS gene had been expressed steadily in Acacia crassicarpa. The transgenic plants of Acacia crassicarpa were gained in the experiment.