| NbDREB1 ( from Nicotiana Benthamiana ) and BnDREB1-5 transcription factor genes (form Canola) were transferred into flue-cured tobacco K326 and oriental tobacco Izmir and Basma respectively using the agrobacterium-mediated leaf discs method. Factors affecting the transformation of tobacco by Agrobacterium tumefaciens were studied and the optimum transformation condition was defined. Transgenic tobacco plants were obtained and their abiotic characteristic was analysed. Now the major results are abstracted as follows: 1.In the process of Agrobacterium infection, there are many factors affecting transformation efficiency. The age of leaf, the position of leaf, the pretreatment, the concentration of Agrobacterium tumefaciens and the time of dark culture were studied. The results showed 20d age of leaf, mid part of the leaf, 64h pretreatment, 0.45 concentration of Agrobacterium tumefaciens (OD600) and in the dark for 3d was the best condition, and had high transformation rate.2.Daub (100mg/L)Km 10μl spot on the leaf of transgenic plant and wild-type plant, observed the color of the spot after 9d.The results showed that the leaf color of wild-type plant turned yellow in the Km spot, but the transgenic plant had not this change. PCR analysis showed 37 plants is positive. 35S: NbDREB1 (K326) 5 plants, rd29A: NbDREB1 (K326) 5 plants, rd29A: BnDREB1-5 (Izmir) 14 plants, 35S: (Izmir) 10 plants, 35S: BnDREB1-5 (Basma) 3 plants, PCR positive ratio is 54%. 3.The leaves of Basma with 35S: BnDREB1-5 transformation presented a better holding water capacity. The density and opening of upper epidermis stoma of transgenic tobacco were obviously lower than wild-type plants, and the relative membrane permeability of wild-type plant leaves was 1.4 times as large as transgenic leaves, the SOD enzyme activity of transgenic tobacco was higher than wild-types, and the rate of water recovery of transgenic Basma was quicker too. After drought... |
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