Study On Genetic Transformation Of Gmftsh Gene In Soybean

Protein FtsH, one of the key components for the plants resisting to photoinhibition and repairing the norm function of PSII, involves in the degradation of PSII core protein D1’s photooxidative damage. Gene GmftsH, cloned from soybean Kefeng1’s No.15chromosome, was transferred into cultivated soybean using pollen tube pathway method and Agrobacterium mediated cotyledon nodes transformation method in order to establish the basis for the cultivation of transgenic soybean varieties with high photosynthetic efficiency.The main results are as follows:1. Gene GmftsH was transferred into7,550flowers from five species of cultivated soybeans using the pollen tube pathway method.3,132pods and6,479seeds of the first generation T1were obtained with an average pods rate of41.36%.34strains of resistant plants were identified using the hygromycin excised leaves detection method,12of which were proved to be inducted with the target gene by PCR detection, with an average conversion rate of0.19%.2. Gene GmftsH was transferred into soybean using the Agrobacterium mediated cotyledon nodes transformation method.4strains of hygromycin resistance plants were obtained,3of which were proved to be inducted with the exogenous gene by PCR detection.3. The hygromycin excised leaves detection method was established for large-scale screening of transgenic soybean, which had the advantages of intuitive results, less work, short cycle and so on. This method was operated as follows:A soybean leaf was cut along the main veins into two pieces, made with3-4rips and placed them in20mgL"1hygromycin testing solution within a6-well cell culture plate. These testing samples were cultivated for16h in light and8h in darkness repeatedly under26℃. After three days, we observed the change of the leaves. The samples with green leaves and incision without browning were resistant leaves. The samples with yellow leaves and browning incision were non resistant leaves.4. The conditions for seeds sterilization and the cultivation time for the Agrobacterium, using the Agrobacterium-mediated cotyledonary nodes transformation method, were both optimized. The results indicated that the best sterilization time for dry chlorine sterilization was12h, which ensured not only the sterilizing effect but also the germination rate of soybean. The optimal cultivation time for Agrobacterium was16～18h, and an appropriate concentration was achieved with the strongest vitality during this time.