1a. Objectives (from AD-416)
The long-term objective is to develop genetics of plant and seed chemistry useful in developing crops with improved end-use quality. The primary crops studied are barley and maize. The primary targeted end-uses are for feeds, foods, and biofuel production. Over the next five years the main focus will continue to be the genetics of plant and seed phosphorus. There are three specific objectives. The first objective will be to identify and characterize genes perturbed in barley low phytic acid mutants. Second, this project will develop a better understanding of the relationship between seed phosphorus and inositol phosphate chemistry, plant performance, and stress tolerance. The third objective is to identify novel crop genotypes conditioning altered plant or seed phosphorus chemistry or related phenotypes that are of end-use value. This includes the development of genetic tools useful in breeding or engineering crops with normal shoot P but altered levels of seed total P, and the development of genetic approaches useful in engineering reduced shoot total P. This last objective involves developing genotypes that are tolerant to reduced plant total P.
1b. Approach (from AD-416)
For the first objective, the first step is to select target mutant loci as targets. Characterization of phenotypes observed in the barley lpa mutation collection, in combination with chromosomal mapping data, and review of the current knowledge in this field, will be used to identify targets for gene identification. Fine mapping of selected lpa loci will first be conducted. Genomics resources for barley, or comparative mapping data with other species such as rice, will be used to identify candidate genes. Sequencing of candidate genes will confirm if they are in fact the gene perturbed in the target mutant. Definitive proof that a mutant phenotype is the result of a mutation in the identified gene may require additional studies such as “complementation”. For the second objective, to develop a better understanding of relationship between seed phosphorus, inositol phosphate and plant performance such as stress tolerance, the first step would be to complete “transcription profiling”, using microarray analyses, to identify genes and functions impacted in selected genotypes or in response to selected stress treatments such as heat/drought stress. Genes whose expression or function is greatly impacted will be the focus of more targeted study. Their sequences, map position, and gene family relationships will be obtained. Expression profiles and biochemical/physiological function will be determined. For the third objective, two types of approaches will be used to develop genotypes with useful alterations in plant or seed phosphorus chemistry. These are “forward genetics” screens and “reverse genetics” approaches. In “Forward genetics”, screens of various types of mutagenized populations will be conducted to identify mutations that impact plant or seed phosphorus. Populations screened will either be chemically mutagenized, or represent collections of transposon insertions. Mutants with interesting phenotypes such as “reduced seed total P” will then be the subject of in depth follow-up study, including chromosomal mapping and agronomic evaluation. Ultimately the gene perturbed in the mutation will be identified, and near-isogenic lines will be developed for use in agronomic and nutritional studies. In the reverse genetics component, mutations such as “single nucleotide polymorphisms” (SNPs) will be isolated in target genes of interest. For example, many genes are already known to be important to phosphorus uptake and transport, but which specific genes or functions are important to seed total phosphorus is not known. Methods such as TILLING will be used to isolate mutations in target genes, which will then be used to determine the effect of such mutations on plant and seed phosphorus.5/17/07.
3. Progress Report
Work continues on the identification of genes important to phytic acid synthesis and accumulation in seeds. Genetic mapping of barley low phytic acid genes has identified six loci, and work continues on fine mapping and identifying these genes. Two of the barley low phytic acid genes have been shown to result in reduced seed total phosphorus, an important finding. This finding is the subject of further studies. About ¼ of the genetic screens for “low seed total P” mutations have been conducted, including screens of both barley and maize. This initial screening work continues as does follow up studies of new putative mutants. Initial progress has been made in setting up “TILLING” for “single nucleotide polymorphisms” (SNPs) that result in recessive alleles of target genes. The first gene identified as a target is one that encodes a phosphorus transporter believed to be important to seed total phosphorus.