1a.Objectives (from AD-416):
The objectives of the project are to (1) assemble a collection of natural accessions and 2,000 homozygous T-DNA lines; (2) conduct a detailed phenotypic characterization of the collection using a phenomic approach, and (3) begin detailed characterization of a select group of mutants and natural accessions.
1b.Approach (from AD-416):
This project builds upon a collection of insertional mutants (T-DNA lines) and natural accessions of Brachypodium that we have already assembled. We will use PCR to identify homozygous mutants lines from segregating populations. We will then use high-throughput, non-destructive phenotyping (phenomics) to characterize the population of insertional mutants and natural accessions. This analysis is expected to identify variation in biomass, growth rate, water use efficiency, nutrient use efficiency and other traits relevant to bioenergy crop development. Based on this data we will select a small subset of lines for more detailed characterization including molecular complementation analysis to determine if the T-DNA insertion is responsible for the observed phenotype. All phenomic data and homozygous mutant lines will be made freely available to the research community.
Our ongoing collaborative project to extensively phenotype Brachypodium distachyon germplasm is almost finished. This work relates to the parent project by providing experimental resources, homozygous T-DNA tagged mutants, and data, phenotyped populations, that are used to achieve the objectives of the parent project more rapidly. We completed phenotyping a collection of 500 homozygous T-DNA lines and are currently analyzing the data. We selected two T-DNA lines for detailed genetic analysis and have demonstrated that the T-DNA is genetically linked to the observed phenotype. We created DNA constructs containing the wild-type gene and are inserting these constructs into the mutants in order to show complementation. This is the final step to definitively demonstrate that the T-DNA insertion is responsible for the observed phenotype. These efforts are creating a valuable genetic resource that will allow researchers to gain the information necessary to use a predictive approach to create improved crop varieties.