The Research On Establishment Of Efficient Transformation Regeneration System Of Sugar Beet And Genetic Transformation

Sugar beet is one of important sugar crops in the world, which provides approximately One-third of the world’s sugar, also one of the two major sugar crops in China. Occupying an important position in the northern agricultural production.In recent 2-3 years with the development of mechanization of sugar beet planting patterns, plant diseases and insect pests in sugar beet production became apparent. Practice has proved that transgene is the most effective way to solve the crop pests and weeds. Therefore, aiming at the main insect pests and weeds in sugar beet production as the control object, be imperative to carry out the research on transgenic technology of sugar beet, and the sugar beet industry long-term development needs. At present, the genetic transformation of plant genetic engineering technology has basically taken shape, but the industrialization of transgenic beet varieties have not been reported. Sugar beet varieties genetically modified failed to industrialization application is caused by a variety of reasons, the two important reason is it difficult to establish a high efficient genetic transformation of beet regeneration system and failed to find a suitable and efficient genetic transformation method. This study based on five different genotype sugar beet variety of explant regeneration culture, analysis the influencing factors of sugar beet explant regeneration, establishment and optimization of regeneration system of sugar beet. Enzyme identification had built the Bt gene express carrier pSKBt crylAc of sugar beets, and through the method of agrobacterium mediated genetic transformation of sugar beet. Study of agrobacterium mediated transformation the different conditions of sugar beet, to establish beet genetically modified system. To lay the foundation for the industrialization of production of transgenic sugar beet varieties The main results are as follows:1. Five different genotypes sugar beet lines(30407、30427、32449、35250、44321) were used as transgene experiment materials in this study, based on the five kinds of materials of explant regeneration tissue culture research, select high induction rate genotype material (35250). To use 35250 as the material, screen is suitable for the strain of the best differentiation medium:MS+0.5mg/L NAA+1.5mg/L 6-BA; subculture medium: MS+0.5mg/L NAA+1.0mg/L 6-BA; rooting culture medium:MS+1.0mg/L NAA.2. Enzyme identification of beet expression vector containing Bt gene crylAc pSKBt, which proved the accuracy of the carrier. By using Agrobacterium mediated transformation method for conversion of sugar beet. The best condition of agrobacterium infect beet was optimized for:OD600= 0.5-0.7, the optimum infection time 25 min; after resistance screening, finally obtained 24 resistant somaclone.3. Using PCR amplification techniques to detect the resistance of plants,5 strains were identified as Bt gene crylAc positive plantlets.