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| Transformation of UDP-glucose Pyrophosphorylase Gene (UGPase) from Saccharum officinarum into Arabidopsis thaliana and Analysis of Physiological Characteristics of Transgenic Plants |
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Abstract UGPase(UDP-glucose pyrophosphorylase) is one of the enzymes which mainly involves in plant carbohydrate metabolism, and plays an important role in the process of plant growth and development. The plant expression vector was constructed by ligating the cDNA fragment of UGPase from Saccharum Officinarum into expression vector pBI121, then the recombinant plasmid pBI-UGP was confirmed by restriction enzyme digestion analysis with BamHⅠ/SacⅠ, and further verified by DNA sequencing. The expression vector was introduced into Arabidopsis thaliana by floral-dip method, with Agrobacterium tumefaciens as media. Five T0 transgenic plants were selected by combining kanamycin screening with PCR detection. After selecting by kanamycin screening, T1 transgenic plants were identified by PCR and Southern blot, the results showed that the target gene was transformed into Arabidopsis thaliana successfully, and different transgenic plants contained various copy numbers of UGPase from Saccharum officinarum. T2 transgenic plants were identified by PCR and RT-PCR, the results indicated that the gene not only inherited stably in self-progeny, but also expressed at the RNA level. Meanwhile, content of soluble sugar, sucrose and starch were determinated in T2 transgenic plants and wild plants. Compared to wild plants, the soluble sugar content of transgenic plants had little change, however, sucrose content of transgenic plants improved obviously, increased by 50.85%~96.99%. Starch content of transgenic plants was lower than that of wild plants, decreased by 9.69%~36.76%. The research proved that UGPase plays a critical role in the process of converting sugar and starch, and the direction of its catalysis influneces distribution of these products in plant organization. It will lay foundation for further study of UGPase.
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Received: 10 October 2011
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