Construction Of Brachypodium Distachyon T-DNA Insertional Mutant Pool

Brachypodium distachyonis, a temperate grass of Poaceae, belonging to the subfamily Pooideae, has been regarded as a suitable model plant for these economically important crops, such as wheat, barley, and so on due to its small stature, the ability to self-pollinate, short lifecycle, simple growth require ments, and small genome size and transformable easily. As genome sequencing and annotation of Brachypodium distachyonis were finished and some new advances have been made. The study of gene function has become the keystone. The most direct and effective way of studying gene function is to construct Brachypodium distachyon mutant pool. T-DNA insertion is one of important methods of construvting mutant pool. The accession BD21, which genome sequense has been completed, was applied to create large collections of sequence-indexed T-DNA mutants. Then, it should play an important role on the closely relationship crops, such as wheat, corn, rice, grasses, turf grass. The main content just as follows:1. Optimization of high frequent regeneration system of Brachypodium distachyonThe effect of different age of callus on the regeneration of embryogenic callus was studied. They were88.6%,87.6%,74.2%for the accessions BD21,BD21-3, and BD018respectively. BDR018gave the highest regeneration ability compared with the other two accessions.2. Establishment of efficient genetic transformation system of Brachypodium distachyonThe accession BD018combined with four different strains of AGLO, AGL1, EHA105, LBA4404(all with binary vector pDM805-containing the Actl:uidA:rbcS reporter gene and the Ubil:bar:nos selectable marker gene) was compared for producing transgenic Brachypodium plants. The results showed that AGL1appeared to be the best for gene delivery. Transformation efficiency was strongly influenced by the selectable marker gene, selection procedure and genotype. The highest transformation efficiency was observed using the hpt gene in combination with hygomycin.3Highly efficient Agrobacterium tumefaciens-mediated transformation methods have been developed for BD21. More than3000T-DNA lines have been created, with the ultimate goal to creat anther15000lines.