Study On The Transformation Of Populus×euramericana Cv. "74/76" With CMO Gene

Poplar is one of the wide-spread cultivated trees in our country. High growth rate, easy propagation and its usefulness in timber industry, pulpwood making, shelter belt and other purpose afforestation make it to be the most important tree for artificial silviculture and for the same reason poplar will be the main tree species to establish million-hectare forest bases for fast-growing commercial wood.Firstly, the optimal media of populusxeuramericana cv."74/76" for bud differentiation and rooting were established. The high frequency of shoot regeneration can be induced on MS medium supplemented with 0.8 mg/L BA, 0.05 mg/L NAA. When the shoots grew to 2-3 cm high, cut them from leaves, and then transferred on to root differentiation medium (MS+0.3 mg/L NAA). The shoots would develop into whole plants within 10 days.The sensitivity of explants to cefotaxine, kanamycin and hygromycin were tested separatly in order to decide a reasonable selective pressure for transformation. Cefotaxine 250 mg/L could inhibit Agrobacterium tumefacien growing too rapidly. Kanamycin 15 mg/L, hygromycin 5 mg/L can be used as the died line concentration of selecting the transgenic bud during the bud differentiation. In the rooting phase, kanamycin concentration shoud be 20 mg/L.The results confirmed that adding 200μmol/L AS into Agrobacterium culture medium could increase Kan-resistant shoots rate.6 kanamycin-resistant rooted plants were obtained through successive selection in shoot and root induction stage under high level kanamycin pressure. PCR and Southern analysis showed that 1 of 6 kanamycin-resistant rooted plants were positive. This meant that CMO gene was successfully integrated into the genome of the plant.The relative electronic conductivity of the transgenic plants under 0.5% NaCl stress for 5 days was about 17.2%, less than the control plants relative electronic conductivity(26.4%), these showed the transgenic plants has been less injured than control plants under salt stress. Salt tolerance test showed that transgenic plants grew better than non-transgenic plant on medium containing 0.4%-0.8%NaCl.