The effect of co-cultivation and selection parameters on <Emphasis Type="Italic">Agrobacterium</Emphasis>-mediated transformation of Chinese soybean varieties |
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Authors: | Sheng-Jun Liu Zhi-Ming Wei Jian-Qiu Huang |
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Institution: | (1) Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, 300 Fenglin Road, 200032 Shanghai, China;(2) Graduate School of the Chinese Academy of Sciences, 300 Fenglin Road, 200032 Shanghai, China |
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Abstract: | In the present study, an efficient Agrobacterium-mediated gene transformation system was developed for soybean Glycine max (L.) Merrill] based on the examinations of several factors affecting plant transformation efficiency. Increased transformation
efficiencies were obtained when the soybean cotyledonary node were inoculated with the Agrobacterium inoculum added with 0.02% (v/v) surfactant (Silwet L-77). The applications of Silwet L-77 (0.02%) during infection and l-cysteine (600 mg l−1) during co-cultivation resulted in more significantly improved transformation efficiency than each of the two factors alone.
The optimized temperature for infected explant co-cultivation was 22°C. Regenerated transgenic shoots were selected and produced
more efficiently with the modified selection scheme (initiation on shoot induction medium without hygromycin for 7 days, with
3 mg l−1 hygromycin for 10 days, 5 mg l−1 hygromycin for another 10 days, and elongation on shoot elongation medium with 8 mg l−1 hygromycin). Using the optimized system, we obtained 145 morphologically normal and fertile independent transgenic plants
in five important Chinese soybean varieties. The transformation efficacies ranged from 3.8 to 11.7%. Stable integration, expression
and inheritance of the transgenes were confirmed by molecular and genetic analysis. T1 plants were analyzed and transmission of transgenes to the T1 generation in a Mendelian fashion was verified. This optimized transformation system should be employed for efficient Agrobacterium-mediated soybean gene transformation. |
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Keywords: | Agrobacterium tumefaciens Glycine max cotyledonary node Transformation |
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