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United States Department of Agriculture

Agricultural Research Service

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Location: Houston, Texas

2011 Annual Report

1a. Objectives (from AD-416)
Objective 1. Use genetic, molecular, and physiological approaches to define the role of specific genes and gene products in the acquisition and whole-organism partitioning of minerals (iron, zinc, calcium, and magnesium) and other factors that inhibit or promote absorption of these minerals in plant foods. Sub-objective 1.A. Identify quantitative trait loci (QTLs) associated with elevated seed or leaf mineral concentrations or seed biomass accumulation. Sub-objective 1.B. Identify rate-limiting and/or novel genes that contribute to tissue Fe, Zn, Ca, and Mg concentrations or seed biomass accumulation. Sub-objective 1.C. Assess the functional role of newly identified gene products in whole-plant nutrition and nutrient partitioning. Sub-objective 1.D. Measure the effects of modulating cation transporter functions on plant Ca, Mg, Fe and Zn content. Sub-objective 1.E. Measure the impact of altered transport function in agriculturally important crops. Sub-objective 1.F. Identify and isolate genes that are involved in calcium oxalate crystal formation in selected mutants. Sub-objective 1.G. Assess the role of the identified genes in calcium oxalate crystal formation. Objective 2. Conduct animal and human feeding studies to determine mineral bioavailability of the nutritionally enhanced crops. Sub-objective 2.A. Use novel transgenic plants to dissect the relationship between mineral partitioning and nutrient absorption in mice feeding studies. Sub-objective 2.B. If the rodent studies demonstrate proof of concept, initiate pilot feeding studies using young adults. Objective 3. Develop new, cost-effective methods for the intrinsic labeling of plant foods for use in nutrient bioavailability studies.

1b. Approach (from AD-416)
The long-term objective of this project is to contribute to the development of nutritionally enhanced plant foods and to develop tools for testing nutrient bioavailability. We have chosen to work initially with plants that are tractable molecular genetic systems (Arabidopsis, Medicago, and soybean) where we can perform gene discovery quickly. We then translate these findings into agriculturally important crops that can be easily transformed and for which established protocols are in place for measuring nutrient absorption in both mice and humans. Specifically, we will work to identify and characterize genes and gene products that are involved in mineral transport throughout the plant, focusing both on whole organ accumulation and subcellular partitioning of minerals. We also will identify and characterize the molecular processes associated with calcium oxalate formation in plants. We envision this work to eventually have relevance to mineral nutrition improvement (e.g., calcium, magnesium, iron, and zinc) in several agronomic crops. In addition, we will develop new, cost-effective methods for the intrinsic labeling of plant foods, using stable isotopes, in order to facilitate nutrient bioavailability studies in humans. These efforts will expand our capabilities for assessing the absorption and metabolism of various plant-derived minerals and phytochemicals. They also will facilitate the generation of new bioavailability data for various nutrients, which will allow informed decisions when policymakers establish future dietary recommendations for humans.

3. Progress Report
To understand the molecular mechanisms regulating calcium oxalate crystal formation in plants, we have completed genetic and molecular studies directed toward identifying new genes required for crystal accumulation in Medicago truncatula. We have completed three back-crossings of two calcium oxalate crystal insertion mutants to allow easier identification of the defective genes. Using back-crossed mutant 1, we have been able to isolate a small fragment of genomic DNA that contains an insertion in mutant gene 1. We are currently using this small fragment of DNA to screen a gene library to isolate mutant gene 1. We have also completed mineral analysis on both back-crossed mutants and determined that their mineral composition is similar to control plants. We have also continued our work to understand how plant calcium transporters have evolved. We have used yeast genetic manipulations and carefully designed experiments in an attempt to "evolve" plant transporters with new properties. We have used biochemical and genetic tools to show that a particular group of transporters work in tandem to perform specific functions. The concept of these transporters working in teams is novel. The knowledge obtained from this work will allow us to genetically modify various plants to alter nutritional content. We have been using quantitative genetics approaches to understand which regions of the plant genome (DNA) are associated with various nutritional traits. We have used unique populations of soybean, bean, or other model plant species to study seed mineral concentrations and root processes that help the plant absorb nutrients, such as iron. We have initiated the seed mineral analysis of a large population of soybeans, grown in replicated field plots in Beltsville, MD. We have also grown several plant species in the CNRC greenhouse or growth chambers in order to assess root traits, leaf mineral, or seed mineral characteristics. These data have been analyzed to identify regions of the plant genome that are linked to specific seed or leaf mineral concentrations, or enhanced ability of the root system to absorb iron. Parent lines have now been crossed with unique genetic lines, in order to produce progeny with an altered genetic makeup. These lines will be used to test and confirm which regions of the DNA are most associated with the measured nutritional traits. The ADODR monitors project activities by visits, review of purchases of equipment, review of ARS-funded foreign travel, and review of ARS funds provided through the SCA.

4. Accomplishments
1. Identifying new genes for enhancing calcium bioavailability in edible plants. Calcium, when present as the calcium oxalate crystal in foods, is unavailable for nutritional absorption. Such crystals are common in edible plant foods, thereby reducing their nutritional quality. Researchers at the Children's Nutrition Research Center in Houston, TX, have identified a small region of DNA within the genome of the model legume Medicago truncatula that contains a previously unidentified gene required for calcium oxalate crystal production. When this gene is "turned off" there is a dramatic reduction in the plants ability to produce these crystals. It is anticipated that the isolation of such a key gene will provide the molecular target that, when inactivated, will enhance the nutritional value of economically important crop plants.

2. Improved yield and nutritional content in melons. Grafting is a method of plant propagation where the tissues of one plant are encouraged to fuse with those of another. It is commonly used for the propagation of many melons grown commercially. Plant scientists at the Children's Nutrition Research Center have successfully used genetically modified rootstocks for grafting of melons. These modified rootstocks produced larger, more robust melons when compared with typical plants. Using genetically engineered rootstocks could be a means of boosting plant productivity for many commercial crops that use grafting techniques. This technique would be particularly compelling for the general public in that the genetic modifications do not enter the food supply.

3. Plant root responds to differing iron deficiency conditions. Plants acquire iron from soils via processes functioning in their roots, but the availability of iron in different soils can sometimes make it difficult for those roots to absorb enough iron to meet their needs. Soils that are alkaline (high pH) and contain high levels of calcium carbonate (also known as calcareous soils) are poor sources of iron, but some plants have found ways to acquire iron in these very challenging soils. Plant scientists at the Children's Nutrition Research Center have found that the roots of a certain legume plant could synthesize and release compounds that increased the levels of available iron in the soil. Moreover these plant roots could also change their internal biochemical properties to help them function more effectively with less iron. The identification of these changes and the identification of some of the genes responsible for them are providing tools and insights to help us develop new crops with improved abilities to acquire iron.

Last Modified: 2/23/2016
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