Location: Plant Science Research2008 Annual Report
1a. Objectives (from AD-416)
Objective 1: Develop germplasm and determine genetic and biological processes that regulate forage use for bioenergy and livestock production. Sub-objective 1.1. Identify genes and breeding strategies to be used for alfalfa improvement. Sub-objective 1.2. Improve energy availability from forages by modifying genetic, metabolic, and developmental processes that control cell wall synthesis and breakdown. Sub-objective 1.3. Identify and utilize mechanisms to improve nutrient uptake in Medicago spp. Objective 2: Develop and evaluate crop management strategies to increase use of perennial forages for livestock and bioenergy, and to protect the environment. Sub-objective 2.1. Develop management practices and systems to optimize alfalfa composition and biomass yield for the efficient production of liquid fuels and syngas. Sub-objective 2.2. Evaluate strategies to reduce root and foliar disease in alfalfa. Sub-objective 2.3. Develop and test management strategies to expand the role of alfalfa and other perennial forages in protecting water quality.
1b. Approach (from AD-416)
Alfalfa is the backbone of sustainable practices for producing crops and animals while protecting water and soil resources. However, use of alfalfa is not always maximized due to several limitations, and new germplasm and management systems are needed for biofuel production. The objectives of this project are to: (1) develop germplasm and determine genetic and biological processes that regulate forage use for bioenergy and livestock production, and (2) develop and evaluate crop management strategies to increase use of perennial forages for livestock and bioenergy, and to protect the environment. Alfalfa germplasm with greater stem in vitro neutral detergent fiber digestibility (IVNDFD) will be developed through selection and breeding utilizing near-infrared reflectance spectroscopy. Populations will be assessed for changes in cell wall chemistry and biofuel conversion under conventional and biomass management systems. Breeding strategies will be evaluated to increase yield potential. Yield will be evaluated in replicated field trials in multiple locations. Populations with highest yield will be evaluated for total forage yield under conventional and biomass management systems. Management methods for reducing diseases that impact yield and persistence will be assessed. The effect of glyphosate on foliar diseases (rust, powdery mildew, anthracnose, spring black stem) will be evaluated in Roundup Ready alfalfa. Alfalfa cultivars will be screened for resistance to brown root rot and the role of crop debris in pathogen survival and increase will be measured. The potential for green manures and traffic tolerance to reduce crown rot will be evaluated. Genes for disease resistance, cell wall biosynthesis, and nutrient uptake will be isolated to provide new knowledge on the genetic underpinnings of these traits and markers for plant improvement. Transcript profiling will be done to identify candidate genes involved in these traits. The function of specific genes will be investigated through detailed transcript expression studies, investigating promoter activity, biochemical assays, over-expression, and gene knock down. Alfalfa germplasm with greater capacity to remove soil nitrate will be developed by recurrent selection using bromide as a nitrate analog. Populations will be evaluated in multiple field locations for bromide and nitrate uptake. Removal of nitrogen from soil by alfalfa will be tested at the field scale in replicated plots in multiple locations. Forage samples will be analyzed for dry matter and N yield; topsoil will be measured for total N and C. Removal of nitrate from irrigation water via phytofiltration will be tested on a field scale. Replicated plots will be irrigated at normal, high, and intermediate rates and the total nitrate leached determined. Biocurtain strips along tile drainage sites will be managed using conventional and biomass systems along side a corn-soybean rotation. Inorganic N will be measured throughout the growing season in tile drainage water. Fulfilling the objectives will help meet the emerging needs of the nation in livestock and bioenergy production and environmental protection.
3. Progress Report
Progress in developing germplasm and determining genetic and biological processes that regulate forage use for bioenergy and livestock production include the following. Final samples for the last cycle of selection for alfalfa stem in vitro neutral detergent fiber digestibility were collected and third cycle populations made. Crosses were completed to create about 150 hybrid and 150 synthetic alfalfa genetic sources to compare selection methods for increasing herbage yield. All of the alfalfa genetic sources were established in replicated field evaluation trials in MN and WI. Seed production for the final cycle of selection for divergent nitrate-N uptake was completed. Field evaluation trial comparing nitrate-N uptake capacity will be established in spring 2009. Completed transcript profiling of roots undergoing phosphorus- (P) and nitrogen- (N) deficiency stress and identified differentially expressed genes. Completed transcript profiling from stems of 2 alfalfa lines differing in cellulose and lignin content to identify candidate genes for further study. Mapman analysis has been applied to all transcript profiling experiments. Completed generation of alfalfa RNAi transformants for key genes regulating cell wall composition. Gene expression in the transformants is being evaluated to identify lines for further study. Transgenic Medicago truncatula (Mt) plants expressing RNAi constructs for 3 candidate genes involved in disease resistance were characterized. Two genes appear to have little effect on disease resistance when down-regulated; down-regulating the third dramatically increases susceptibility. Two different genes putatively down-regulated in alfalfa also resulted in increased susceptibility. In support of research to identify a major gene controlling powdery mildew resistance, 228 Mt recombinant inbred lines were screened for disease phenotype. Progress in developing and evaluating crop management strategies to increase use of perennial forages for livestock and bioenergy include the following. Laboratory analysis of cell wall composition and digestibility for biomass-type alfalfa stem samples was completed, the near-infrared reflectance prediction equations were updated and calibrations used to select 92 alfalfa stem samples that represented the diversity available for this candidate bioenergy crop. The calibration sample set was sent to a collaborator to determine cellulosic ethanol conversion potential. From the alfalfa clones previously retained with divergent stem cell wall composition, the most extreme clones for divergent cellulose and lignin concentrations were selected for vegetative propagation and initiation of a cellulosic ethanol efficiency trial beginning in FY 2009. A set of 20 alfalfa stem samples that spanned the range in lignin concentration observed in this crop was subjected to pyrolysis as a measure of potential thermochemical conversion to syngas in collaboration with a lab in PA. Roundup-Ready alfalfa plants were evaluated for resistance to rust and powdery mildew. Glyphosate treatment was found to protect plants from rust infection in greenhouse experiments. (NP 215, Component 3, Objectives H1, H2, J1, J2)
1. Pyrolysis Product Yields for Bioenergy are not Significantly Impacted by the Natural Range in Lignin Concentration of Alfalfa Stems Gasification is a modified combustion process that can create both syngas for liquid fuel production and heat for electricity generation. The two major components of biomass are carbohydrates and lignin, with lignin having almost twice the raw energy content of carbohydrate. Pyrolysis, a process integral to gasification, was used to compare a set of alfalfa stem samples that spanned the natural range in lignin of this crop to determine if lignin content would impact gasification response. Although yield of a few specific types of gases from pyrolysis was impacted by alfalfa lignin content, overall yield of the major gas product categories and residual combustible charcoal was not affected by biomass lignin content. This was in stark contrast to the very strong negative impact of lignin on biological fermentation. These results clearly indicate that genetic modification and harvest management of alfalfa to provide stems with less lignin for processing to bioenergy by gasification is not necessary, in contrast to lignin's extreme importance to cellulosic ethanol conversion efficiency. (NP 215, Component 3, Problem Statements H and J)
5. Significant Activities that Support Special Target Populations
Boateng, A.A., Weimer, P.J., Jung, H.G., Lamb, J.F. 2008. Response of thermochemical and biochemical conversion processes to lignin concentration in alfalfa stems. Energy and Fuels. 22:2810-2815.