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United States Department of Agriculture

Agricultural Research Service

Research Project: Enhanced Alfalfa Germplasm and Genomic Resources for Yield, Quality, and Environmental Protection

Location: Plant Science Research

2013 Annual Report


1a.Objectives (from AD-416):
Objective 1: Develop strategies and tools for improving alfalfa yields, disease resistance, and nutrient cycling efficiency. Sub-objective 1.1: Identify and compare breeding strategies for alfalfa yield improvement. Sub-objective 1.2: Identify strategies and develop tools for reducing losses from diseases. Sub-objective 1.3: Develop strategies to produce alfalfa germplasm with improved herbage yield and nutrient cycling (phosphorus (P) and potassium (K)) in pure stands and in mixtures with forage grasses. Sub-objective 1.4: Develop a gene expression atlas for two divergent alfalfa gene pools (Medicago sativa subsp. sativa and M. sativa subsp. falcata), identify organ-specific genes, and mine sequences for gene pool diversity (SNPs and SSRs). Objective 2: Develop forage germplasm with modified cell-wall structure and chemistry to improve digestibility, and evaluate impacts on livestock and bioenergy productivity. Sub-objective 2.1: Assess alfalfa germplasm altered for in vitro neutral detergent fiber digestibility (IVNDFD) for forage yield and energy availability for livestock production and biofuel conversion efficiency. Sub-objective 2.2: Identify genetic, metabolic, and developmental processes in alfalfa stems that regulate cell wall composition and energy availability. Objective 3: Develop alfalfa germplasm and crop rotation management systems to improve nitrogen cycling and carbon sequestration. Sub-objective 3.1: Identify and utilize mechanisms to improve nutrient uptake in alfalfa. Sub-objective 3.2: Identify and characterize rhizosphere soil microbes that promote carbon sequestration and improve the agronomic and environmental benefits from crop rotation. Sub-objective 3.3: Measure and predict N credits for second-year corn grown after alfalfa to improve N management and reduce N losses.


1b.Approach (from AD-416):
To increase yield potential, the contribution of heterosis to yield potential and the effectiveness of selecting for high yield per stem as a yield component in alfalfa will be assessed. Synthetic populations, semi-hybrid populations, parent populations, and two commercial varieties will be established in replicated field trials and total annual forage yield will be evaluated for at least two production years. We will develop DNA markers to increase disease resistance and measure diversity in pathogen populations. A bulked segregant analysis will be done using populations segregating for resistance to Aphanomyces root rot. Histochemical and gene expression studies will be used to gain an understanding of the infection process and mechanisms of resistance in resistant and susceptible plants. Simple sequence repeat (SSR) markers will be identified in the Verticillium albo-atrum genome sequence and tested for polymorphisms on field isolates and standard strains. Plant responses to these strains will be measured with the standard disease severity scale and the amount of pathogen present determined by the qPCR method. Plants with the lowest amount of pathogen present will be retained, intermated, and progeny tested for Verticillium wilt resistance. DNA markers will also be developed from a gene expression atlas for two divergent alfalfa gene pools using transcripts from leaves, roots, nodules, flowers, and elongating and post-elongation stem internodes. To develop alfalfa germplasm with improved herbage yield and nutrient cycling, germplasm differing in root system architecture will be examined in replicated field experiments to determine: P and/or K uptake capacity under low and adequate soil nutrient levels; symbiosis with arbuscular mycorrhizal (AM) fungi; and prevalence of root and foliar diseases. Alfalfa germplasm selected for in vitro neutral detergent fiber digestibility (IVNDFD) and original parents will be evaluated in replicated field trials for forage quality traits, gain from selection, and heritability estimates. Replicated sward plot field trials will be used to determine forage yield and the best crop management methods for germplasm selected for IVNDFD. Energy availability for livestock and biofuel conversion in the harvested forage will be determined by near infrared reflectance spectroscopy. To improve alfalfa stem cellulose content, a comparison of miRNA profiles in elongating stem and post-elongation stem internodes will be used to identify miRNAs that play key roles in the development of secondary xylem. The microbial communities in the rhizosphere that influence plant growth and carbon sequestration will be characterized using culture-dependent and metagenomics approaches. Field tests will determine whether selection for nitrate uptake alters yields of alfalfa-grass mixtures. On-farm field experiments will be established at 10 locations to improve predictions of whether nitrogen contributed by alfalfa to subsequent corn crops will improve farm profit and reduce nitrogen losses.


3.Progress Report:
This project aims to increase alfalfa yields and utilization for livestock and cellulosic biomass and to amplify alfalfa’s environmental services by contributing new knowledge from genes to fields. Progress was made on all project objectives. Under Objective 1 progress was made on developing strategies and tools for improving alfalfa yields, disease resistance, and nutrient cycling efficiency. Field trials to evaluate forage yield potential between hybrid and synthetic populations as well as a populations selected for yield per stem were successfully established in Minnesota and Wisconsin. Initial work to develop tools for improving breeding for disease resistance included establishing a collection of the Verticillium wilt pathogen from symptomatic alfalfa plants, designing primers for SSR markers, and identifying polymorphic markers in 19 strains from the 1980s, which were found to be genetically similar. In research to create a gene expression atlas for alfalfa, 36 libraries were created and used for RNA-seq transcript profiling, sequences have been assembled, and a database compiled. A website for accessing the data is under construction. Under Objective 2 progress in developing forage germplasm with modified cell-wall structure and chemistry to improve livestock and bioenergy productivity was made. Final field samples were collected from experiments evaluating selection for in vitro neutral detergent fiber digestibly (IVNDFD) and its impact on livestock nutrition and biofuel conversion efficiency. New field trials were established to investigate whether a near infrared spectroscopy (NIRS) selection methodology used to create alfalfa populations that differ in stem IVNDFD can be used to develop alfalfa with increased whole forage IVNDFD. Under Objective 3 progress in developing alfalfa germplasm and crop management systems to improve nitrogen cycling and carbon sequestration was accomplished. Progress was made on the question of whether root system architecture of alfalfa influences nutrient uptake or the presence of root or crown disease. Sampling of plants and soils in a three-year field research program was completed at three sites in Minnesota to measure forage production, P and K uptake, and disease incidence. An intensive four-year project was completed to determine the performance of alfalfa selected for high or low nitrate uptake capacity. Final sampling of plants was completed, as were all tissue analyses, and statistical analyses are being conducted. Field experiments were completed on fertilizer N response of second-year corn after alfalfa and a manuscript is being drafted. The key finding is that in about one-half the cases, second-year corn after alfalfa needed no additional fertilizer N to produce the best economic yields. Combined with our earlier findings, this indicates that alfalfa can, under a range of conditions, supply enough nitrogen for two crops of corn, a savings of about $160 per acre.


4.Accomplishments
1. Alfalfa supplies more nitrogen than suspected. Legumes, such as alfalfa, have been used for millennia to provide nitrogen to subsequent crops, but in today’s agriculture, fertilizer nitrogen often is added to guarantee highest economic returns. Although little fertilizer is needed for the first crop of corn after alfalfa, farmers and researchers have little confidence in how much nitrogen alfalfa supplies to a second crop of corn. Collaborative, multi-year, on-farm research between an ARS scientist in Saint Paul, Minnesota, and faculty from the universities of Minnesota and Connecticut produced the largest compilation of research data by combining results from 28 on-farm trials with 39 published experiments. About 50% of the time, alfalfa provides all the nitrogen needed for both the first and second crops of corn, saving about $160 per acre and reducing the amount of fossil fuel needed to produce corn. In other cases, the amount of nitrogen needed for highest yield of the second corn crop varied widely, but in only 10% of the cases did it need the same dose of fertilizer as required by continuous corn. Once scientists learn to predict the fertilizer needs of second-year corn, farmers will be able to save money and reduce the environmental damage caused by over-fertilizing.


Review Publications
Samac, D.A., Lamb, J.F., Kinkel, L.L., Hanson, L.K. 2013. Effect of wheel traffic and green manure treatments on forage yield and crown rot in alfalfa (Medicago sativa). Plant and Soil. DOI:10.1007/s11104-013-1746-5.

Last Modified: 10/21/2014
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