Cloning, Expression Analysis And Genetic Transformation Of GF 14-A-like Gene From Brachypodium Distachyon

Wheat is one of the important food crops. Its research is still a challenge due to the complexity of its genome. Brachypodium distachyon belongs to Pooideae, which has a relatively close kinship with grass crops including wheat. Since B. distachyon has many advantages, such as short growth cycle, small genome, the superiority of easy genetic transformation and complete genome sequencing, scientists can do research at genomic level, which has gradually made it become a new model monpcot plant.14-3-3 is a widespread protein in plants and it can interact with target proteins which has phosphorylation sites, and through forming hetero- or homo-dimer to regulate growth and development of plants. However, there are few reports about the function of 14-3-3 in B.distachyon.In this study, a gene of B.distachyon 14-3-3 family was cloned and named GF14-A-like after sequence alignment. The main results are as follows:(1) After searching in the database, we got tthe full length sequence information of Bd GF14-A-like, and its ORF is 753 bp encoding a peptide of 250 amino acids. The full length c DNA sequence was cloned by nested PCR. After the protein phylogenetic tree analysis among 14-3-3 family members of model organisms rice and Arabidopsis, we found that Bd GF14-A-like shares the highest homology with At GRF1 and Os GF14 a.(2) Transient expression analysis via particle bombardment-mediated onion subcellular localization reveled that Bd GF14-A-like was localized in the nucleus, cytoplasm, and cell membranes.(3) Expression profile analysis of B. distachyon's roots, stems, leaves and fruit showed that Bd GF14-A-like had the lowest expression in roots and the highest in leaves and fruit.(4) After treating the roots and leaves with various abiotic stressed, it was found that expression of Bd GF14-A-like could be upregulated by Na Cl and ETH and downregulated by ABA as reveled by semi-quantitative RT-PCR..(5) We successfully obtained 22 positive transgenic tobacco plants after transformation of plant overexpression vector p BI121-Bd GF14-A-like via Agrobacterium-mediatedgenetic transformation method. We transplanted the seedlings to the greenhouse and successfully got T1 generation.Research on B. distachyon Bd GF14-A-like gene can provide useful imformation for the furture research on the function of 14-3-3 protein in wheat and other important food crops, and for enhancing wheat resistance and improving some of its agronomic traits.