1a. Objectives (from AD-416):
The global objective of this project is to improve productivity and sustainability of cool season food legume production systems in the US. Specific objectives will include: 1. Developing new varieties and germplasm of lentils, peas, and chickpeas that are improved for a range of traits including yield, disease resistance, and cold tolerance; 2. Optimize enabling technologies including molecular markers and transgenic approaches to facilitate variety improvement in cool season food legumes; 3. Determine relative contributions of genetics and environment to variation in peas, lentils, and chickpeas for seed nutritional traits; and 4. Improve nitrogen fixation in peas, lentils, and chickpeas.
1b. Approach (from AD-416):
Cyclical hybridization including single, double and triple crosses will be used to combine favorable alleles for traits of interest. Parental lines will include adapted germplasm, commercial cultivars and germplasm accessions having specific traits of interest such as disease resistance, superior quality, increased biomass production and greater seed yield potential. A modified bulk-pedigree method will be used to advance generations toward homozygosity and identify superior selections. Bulk populations ranging in generation from F2 – F6 for each cross will be evaluated in the field, where populations deficient in easily assayable traits such as height, tolerance to lodging, and early maturity will be identified and dropped from the breeding program. Promising lines will be evaluated over multiple locations in WA and ID for at least 3 years prior to consideration for release as a new variety or adapted germplasm. Methods will be determined for regenerating lentils, peas, and chickpeas from tissue culture. The susceptibility of these crops to various selectable markers including resistance to hygromycin and kanamycin will be determined. Protocols for transforming explants of these crops using Agrobacterium tumefaciens will be developed. Seeds (with seed coats) will be ground to a uniform powder then wet digested and resuspended using trace metal-grade nitric acid and hydrogen peroxide. Digests will be analyzed for mineral concentration using inductively-coupled plasma atomic-emission spectroscopy (ICP-OES). This technique will provide analysis of K, P, Ca, Mg, Fe, Mn, Zn, Cu, Ni, and Mo. Seed protein concentrations will be calculated based on seed nitrogen concentrations, which will be established using a LECO FP-528 Nitrogen/Protein Determinator, using the calculation: seed nitrogen concentration x 5.48 = seed protein concentration. Strains of R. leguminosarum will be isolated from nodules of various lentil and pea cultivars grown in several different locations in WA. Strains of M. ciceri will be similary isolated from nodules of several chickpea cultivars. Genetically distinct isolates of R. leguminosarum and M. ciceri will be identified by using polymerase chain reaction (PCR) to amplify 16S rDNA, followed by sequence analysis. At least five distinct isolates of R. leguminosarum and five distinct isolates of M. ciceri will be used to examine rhizobia genotype effects on biomass and nitrogen fixation in peas, lentils, and chickpeas. Replicated plants will be grown individually in sterile “conetainers” using a low-nutrient sandbased potting mix. Each conetainer will be fertilized with ammonium sulfate with 10% isotopic 15-N and inoculated with a single rhizobium isolate. The number of nodules per plant will be determined. Biomass will be dried, weighed, ground, and analyzed for both total N and 15-N. The proportion and total amount of plant N that was fixed by the rhizobial symbiont will be determined with this information.
3. Progress Report:
Yield trials were conducted at several locations in Washington and Idaho on advanced and preliminary pea, lentil and chickpea breeding lines. Lines were also scored for important production and market traits including plant height, tolerance to lodging, seed size and seed color. Breeder seed is being produced of a promising large seed kabuli line and a “Richlea” market class line of lentil. The results of this project relate to Subobjective 1A (Develop pea, lentil and chickpea cultivars with broad adaptation to diverse production environments and with resistance to fungal and viral pathogens) of the in-house associated project.