Cloning And Functional Analysis Of VRT2-like Genes In Brachypodium Distachyon L.

Plants exhibit a complex regulation of the transition of shoot apical meristems from vegetative to reprodutcive stages to optimize flowering time and seed production. The genes and molecular mechanisms controlling flowering have been extensively studied in the model dicot Arabidopsis thaliana. Emerging evidence suggests that mechanisms of flowering time control are somehow different between monocots and dicots. Here we choose Brachypodium distachyon as a model plant to explore the flowering mechanism typical for cereal grasses such as wheat, barley, oat, and so on.Current evidence suggested that wheat VRT2 is a flowering time gene involving the prevention of flowering in short days. It's reported that VRT2 protein could bind to the promoter of VRN1 to repress its expression, working in concert with VRN2 to prevent flowering. So far little is known on the mechanisms of VRT2 regulating flowering moleculary and biochemically. In this research, two novel BdVRT2-like genes were isolated from Brachypodium distachyon by RT-PCR and RACE-PCR techniques, which are named as BdVRT2-like1 and BdVRT1-like2, respectively. Sequence analysis showed that the BdVRT2-like1 gene contained an open reading frame of 675bp encoding a putative protein of 225 amino acids. And the BdVRT2-like2 gene contains an open reading frame of 690bp encoding a putative protein of 230 amino acids. The molecular characteristic and phylogenetic analysis revealed that BdVRT2-like genes are homologous to MADS-box proteins in the StMADS11-like clade.The expression patterns of BdVRT2-like1 indicated that its transcripts are abundant in shoot apex and in the vegetative stage and low in late reproductive stage. Its expression can be up-regulated by low temperature, but is repressed in a prolonged cold (vernalization), which avoid the transition from vegetative to the reproductive stage during the cold period. Its expression is up-regulated under short-day conditions, but down-regulated under long-day conditions. Over-expression of BdVRT2-like1 in Arabidopsis leads to a late flowering. Which RNAi lines of Brachypodium against BdVRT2-like1 is early flowering. There are some differences between BdVRT2-like2 and BdVRT2-like1. The expression of BdVRT2-like2 is up-regulated gradually in vegetative stage, but its expression is up-regulated in reproduvtive sage. Its expression is higher in shoot apical meristem than other tissue. Its expression is a little bit higher under long-day conditions than in short-days. So photoperiod produces little influence on BdVRT2-like2 expression. These results demonstrated that BdVRT2-like1 may regulate the floral transition, which may be a repressor of the FLoral transition in Brachypodium, which means that BdVRT2-like1 could be the closest homologue of TaVRT-2.