Preliminary Analysis Of The Function Of The Bd FUL1 And Bd FUL2 Genes In Brachypodium Distachyon

Rice, common wheat and maize are all major grain crops in the world.They are closely related to human life. Suitable flowering time and normal flower development are the foundation of seed formation, and further affect the production and quality of grain crops.Brachypodium distachyon is a new model plant of gramineous plant, with its traits: small stature, easy to manage, similar flower organ structure to wheat and rice. More importantly,its genome is relatively small, only 272 Mbp. Therefore, we can study Brachypodium distachyon to solve biological problem of gramineous crops.FUL(FRUITFULL) genes belong to FUL sub-family of MADS-box genes in plants.They take important part in the regulation of flowering time, development of flower organs and seeds, etc. Moreover, their functions are not conservative. In recent years, scientists have made great progress in the studies of FUL gene functions, especially in the model plant Arabidopsis(Arabidopsis thaliana) and rice(Oryza sativa). In this paper, we taken BdFUL1 and BdFUL2 genes as research objects, analyzing the functions of this two genes. On this basis, over-expression and RNA interference vectors were constructed respectively. In addition, we also successfully put BdFUL2 over-expression vector into rice plants by transgenic technology.The research results are shown below:1. In our laboratory, DouYanHua cloned BdFUL1 gene and discovered two alternative splicing transcripts, BdFUL1-1 and BdFUL1-2. On the basis, we also cloned BdFUL2 gene.Then, using the method of bioinformatics, we preliminarily analyzed their sequences and protein structures, constructed phylogentic tree. The results showed that BdFUL1, wheat WFUL1 and rice OsMADS14 affiliated with the same branch. BdFUL2, wheat WFUL2 and rice OsMADS15 affiliated with another branch.In short, the genes that affiliated with the same branch had a close genetic relationship and were likely to have similar functions.2. In our laboratory, DouYanHua preliminarily analyzed expression patterns of BdFUL1-1 and BdFUL1-2 in different organizations. On the basis, we further analysedexpression patterns of BdFUL1-1 and BdFUL1-2 in each round of flower organ. To BdFUL2 gene, we analysed its expression patterns in different organizations and each round of flower organ. Results showed that, unlike BdFUL1 gene, the expression quantity of BdFUL2 in young roots was not high, and the expression quantities in the mature stems and flowers were similar to that of BdFUL1 gene, higher than those in other organizations. Expression quantity analysis in each whorl of flower organ showed that, expression quantities of BdFUL1-1,BdFUL1-2 and BdFUL2 were similar, they all had a high expression quantities in glumes,lemmas, a certain expression quantities in paleas, and also had a low expression quantities in lodicules, stamens, and carpels.3. We analysed expression quantities of BdFUL1 and BdFUL2 in leaves and flowers under the condition of vernalization and unvernalization. Furthermore, we also analysed expression quantities of BdFUL1-1and BdFUL1-2 under the conditions of high temperature,low temperature, drought, salt stress treatments and hormone treatments. Results of vernalization and unvernalization showed that, in the first leaf after heading under the condition of vernalization, BdFUL1-1, BdFUL1-2 and BdFUL2 had higher expression quantities than those in other leaves and flowers. Results of stress treatments and hormone treatments showed that, under the condition of high temperature, low temperature and hormone ABA, 6-BA, SA treatmensts, the expression quantities of BdFUL1-1 and BdFUL1-2increased. Furthermore, the expression quantity of early terminated BdFUL1-1, were significantly higher than that of the complete BdFUL1-2. Under the condition of drought stress(PEG treatment), the expression quantity of BdFUL1- 1 decreased.4. We completed the constructions of three over-expression vectors and four RNA interference vectors for BdFUL1 and BdFUL2 genes respectively. In addition, we also put BdFUL2 over-expression vector into rice plants by transgenic technology successfully. Finally,we analysed expression quantities of BdFUL2 gene in transgenic rices. Results showed that the transgenic rice plants do had the expression of BdFUL2 gene.The above results, sequence analysis, expression pattern analysis, vernalization and unvernalization analysis, high temperature, low temperature, drought, salts tress treatments and hormone treatment analysis, suggested that BdFUL1 and BdFUL2 genes, may be expressed in leaves affected by low temperature vernalization before anthesis, inducing Brachypodium distachyon flowering. BdFUL1 and BdFUL2 genes also regulated external floral organ development. BdFUL1-1 and BdFUL1-2, two alternative splicing transcripts of BdFUL1 gene, may produce under abiotic stress.Their functions were associated with response to environmental stresses. BdFUL1-1 that translates an uncomplete protein, can help Brachypodium distachyon adapt high and low temperature stress and hormonal changes.Construction of over-expression vectors and RNA interference vectors, successful transformation of BdFUL2 over-expression vector in rice laid a solid foundation for further exploration of BdFUL1 and BdFUL2 gene. These findings also provide data for further functional elucidation of regulation of plant flowering time, floral organ and seed development.