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.
3. Progress Report
Progress was made in all three objectives, each of which is important to National Program 301, Component 2, Crop Informatics, Genomics and Genetic Analyses, Problem 2C, Genetic Analyses and Mapping of Important Traits. This project focuses on the genetics of plant and seed phosphorus. Both the total amount of phosphorus and its chemistry are critically important to the end-use quality of grain and legume crops, to the management of phosphorus in agricultural production and to efforts to reduce the impact of agricultural production on the environment. Progress was made on Objective 1, genetic studies of low phytic acid mutants and genes. Genetic tests have now identified five barley low phytic acid genes, and work continues on identifying allelic relationships, fine mapping and the ultimate identification of these genes. Progress was made in Objective 2, which focuses on the relationship between seed phosphorus and inositol phosphates and plant performance and stress tolerance. Via the evaluation of near-isogenic lines, we have identified one barley lpa gene which enhances seed nutritional quality but which appears to have little or no impact on plant yield or stress tolerance. Also, a new class of compounds, the inositol pyrophosphates, important to metabolic-status sensing and stress tolerance, were identified for the first time in plant tissues. Finally, substantial progress was made in Objective 3, which focuses on developing new crop genotypes with altered plant or seed total phosphorus. Screening methods useful in identifying “seed total P” mutants were evaluated and optimized and are currently in use. A gene which this project previously demonstrated to be important to seed total P has now been shown to be a “filial” determinant of seed P uptake. Finally, a new collection of maize mutants which impact seed total P and seed is currently being analyzed in further genetic and chemical detail.
1. Elucidation of an important component of the genetic regulation of seed total phosphorus. A potentially valuable strategy to reduce the environmental impact of dairy and beef production and other forms of animal agriculture, would be to minimize the amount of nutrients in feeds to a level that both maintains productivity and results in reduced waste. A key concern in this area is the typically higher than necessary levels of feed phosphorus and the resulting animal waste phosphorus. As a result breeding “low total P” crops is an important goal but to do so we need to learn more about the genetic control of plant and seed phosphorus levels. As part of this effort in 2010, ARS researchers in Aberdeen, ID, demonstrated that one function of the barley lpa1 gene is to generate a signal in the developing seed that regulates the amount of phosphorus transported into it from the mother plant. This explains why mutations in this gene results in levels of reduced seed total P that would be adequate for the development of environmentally-friendly feeds.
2. Identification of metabolic sensors in plants that help to regulate stress response. Enhancing stress-tolerance is critical to enhancing crop productivity, especially at a time when drought and heat stress may be increasing. In this area of work, we documented for the first time that plant cells synthesize a class of compounds called inositol pyrophosphates. These compounds function as molecular signals that sense plants metabolic status and thus help to regulate how they respond to stress. In 2010, ARS researchers in Aberdeen, ID, demonstrated that mutations in a gene in maize (lpa1) result in accumulations of an inositol pyrophosphate called, inositol octaphosphate (Ins P8), proving that this and related compounds can be synthesized by plants and that maize lpa1 lines represent a good source of such compounds.