Studies On The Transformation Of1-SST Genes Into Cassava

In this study, the genetic transformation systems of several improved cassava cultivar in China is optimizeed. Meanwhile, we detailed important factors of the regeneration efficiency in seven different kinds of Cassava (SC6, SC7, SC8, SC124, NZ188, C3and C4). And agrobacterium mediated transformation system of SC6and SC8cassavas optimization was also optimized on this basis. We also make a study on the Genetic transformation of sucrose fructosyl-transferase gene1-SST in transformated cassava. The positive plant was verified by PCR, RT-PCR and Southern Blot. The main findings are as follows:1. Several significant factors were studied,which impact the regeneration system of cassava. Varieties of SC6, SC7, SC8, SC124, NZ188, C3and C4were used as experimental materials. The results showed that:6-BA had a significant inhibitory effect on apical dominance of cassava, the higher the concentration the stronger the inhibition;2,4-D and picloram had almost equal effect on promoting Somatic embryogenesis. In the process of induction of primary somatic embryos, embryogenic rate of all breeds reached100%, except of SC7, which remained a low embryogenic rate of Secondary somatic embryos, embryogenic rate of Secondary somatic embryos of other varieties grow up to90%; The ripening time of somatic embryos had a great influence on the organogenesis of cotyledon. Frequency of cotyledon organogenesis was high, which is around2weeks after embryo maturation and the induction frequency could be more than70%; The somatic embryogenesis ratio and cotyledon organogenesis frequencies were very different among distinct genotypes, SC8and NZ188cultivar had the highest and second-highest frequency respectively, SC7cultivar displayed the lowest frequency. As an additional content AgNO3(5mg/L) in subculture could help with improving the ability of organogenesis capacity of mature cotyledonary.2. Cassava Agrobacterium-mediated transformation system has been optimized, the best infective conditions are as follows:(1) OD600of bacteria liquid concentration is1.0,(2) Additive amount of AS=150μM,(3) the time of infecting is45min (4) time for co-culturing is3d. In order to ensure the success of the conversion and avoid blocking of plant regeneration process, we gradiently add antibiotics in the medium in the screening and the regenerating process.3. We got71and83PCR positive transformed plants through the optimization of genetic transformation system of SC6and SC8varieties of cassava transformation of1-Sst genes. Among them, there are2RT-PCR-positive transgenic plants and one SC6Southern Blot-positive transgenic lines. Southern Blot analysis of SC8is in progress. These results proved the feasibility of the optimization system.