2013 Annual Report
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.
In 2013, advanced yield trials for peas, lentils, and chickpeas were planted at several locations in Washington and Idaho. Breeding lines, commercial varieties, and populations are being evaluated in field nurseries for reaction to several diseases including Ascochyta blight, Fusarium wilt, and Aphanomyces root rot. Chickpea populations are being examined with DNA markers in the laboratory and evaluated in the field for important traits including disease resistance, maturity, yield, and seed size.