Cloning And Genetic Transformation Of The Multiprotein Bridging Factor 1, A Transcriptional Co-activitor Of Soybean

Adversity of stresses such as drought, cold, salinity, flood and so on are the important factors that limit plant growth and regional distribution and seriously affect crop yield and quality. Transcriptional factors can regulate plant response to adverse stresses, transcriptional coactivators can enhance the combination of transcriptional factors and cis-acting element.MBF1, multiprotein bridging factor 1, is known to be a transcriptional coactivator that mediates transcriptional activation by bridging between an activator and a TATA-box binding protein. MBF1 was first purified from posterior silk gland extracts of silk-worm, Bombyx mori, lately, it was found in plants. It was reported that the expression of MBF1 in Arabidopsis thaliana can enhance the ability of plants to resist adverse stresses, but the function of MBF1 in soybean has not be clear yet. We cloned the soybean MBF1 genes and plan to overexpress the genes in Brachypodium distachyon to provide foundation for functional analysis of MBF1 in soybean. The main results are as follows:1. The MBF1 genes have been cloned and analyzed in soybeanThrough sequence BLAST search and analysis, three soybean MBF1 homologous genes were obtained, including GmMBF1-1 、 GmMBF1-2 and GmMBF1-3, we cloned this three genes from soybean by RT-PCR. The amino acid sequence similarity of the three genes was99% and with similarity of 80% compared to amino acid sequence of Arabidopsis AtMBF1 a and AtMBF1 b. In the phylogenetic analysis, three MBF1 homologous genes were clustered in the same branch. Gene structure analysis showed that three genes were highly coserved in the number and length of exons and introns.2. The expression feature of MBF1 was analyzed by semi-quantitative RT-PCRWe analyzed the expression features of MBF1 in soybean leaves by semi-quantitative RT-PCR with soybean Tubulin gene as internal control, the results showed that the expression of GmMBF1 s may be induced by NaCl、SA and cold, GmMBF1-1 and GmMBF1-2 may be induced by ABA.3. The overexpression vector PC186/GmMBF1-1 、 PC186/GmMBF1-2 and PC186/GmMBF1-3 have been constructedThe three MBF1 homologous genes were ligated to PC414 C vector that digested by NotⅠ and Asc Ⅰ to construct the plasmid PC414C/GmMBF1-1 、 PC414C/GmMBF1-2 and PC414C/GmMBF1-3. Then the gene fragments ligated to PC186 vector by LR restructing reaction. In this way, we constructed three overexpression vector PC186/GmMBF1-1 、PC186/GmMBF1-2 and PC186/GmMBF1-3.4. Gained the transgeneic Brachypodium distachyonWe induced Brachypodium distachyon callus tissue from embryo of seeds, the MBF1 homologous genes were transformed into callus tissue by Agrobacterium GV3101, then we got seedlings by regeneration and rooting of callus.5. The phenotype of transgenic Brachypodium distachyonThe GmMBF1-1 and GmMBF1-3 transgenic T0 generation plants have showed that flowering time delayed compared with common plants, whereas the phenotype of GmMBF1-2transgenic plants have not showed significant change.