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ARS Home » Plains Area » Lincoln, Nebraska » Wheat, Sorghum and Forage Research » Research » Research Project #424223

Research Project: Genetic Improvement of Sorghum for Non-Grain Energy Uses

Location: Wheat, Sorghum and Forage Research

2018 Annual Report

1a. Objectives (from AD-416):
The long-term objectives of this project are to develop improved sorghum (Sorghum bicolor) germplasm (Figure 2) for biofuels production by 1) determining genetic, biochemical, and physiological mechanisms controlling traits beneficial to bioenergy conversion technologies (saccharification & fermentation, pyrolysis and combustion), 2) develop and release improved germplasm with these modified traits, and 3) determine the impact of fungal pathogens on sorghum with these modified traits. Over the next five years, the following specific objectives will be addressed: Objective 1: Determine and manipulate the genetic, biochemical and physiological mechanisms controlling the biological pathways involved in non-grain energy sorghum germplasm. Sub-objective 1.A: Determine the effects of six newly identified brown midrib (bmr) mutants on lignin synthesis and the monolignol biosynthetic pathway. Sub-objective 1.B: Develop transgenic lines over-expressing genes in monolignol biosynthesis to determine their impact on lignin content and composition. Sub-objective 1.C: Identify strategies for increasing the sugar content of sweet sorghum juice and improving its biomass composition for thermal conversion. Objective 2: Determine the impact of fungal pathogens on non-grain energy sorghum germplasm, and determine mechanisms of resistance to sorghum pathogens. Sub-objective 2.A: Determine the response of sorghum with modified lignin biosynthesis pathways to stalk pathogens. Sub-objective 2.B: Determine whether modifications in the lignin biosynthetic pathway affect growth of the pathogen Colletotrichum sublineolum within sorghum leaves. Sub-objective 2.C: Identify sweet sorghum parental lines with resistance to stalk pathogens. Objective 3: Develop and evaluate germplasm to improve sorghum for non-grain energy uses. Sub-objective 3.A: Determine the effects of brown midrib (low lignin) mutations alone or in combination on bioenergy conversion via saccharification and fermentation. Sub-objective 3.B: Develop sorghum lines with novel lignin composition and determine the effects on bioenergy conversion. Sub-objective 3.C: Develop sweet sorghum germplasm incorporating bmr6 and bmr12 to reduce lignin and determine the effects of these alleles on end-use quality.

1b. Approach (from AD-416):
The overall objective of this project is to improve sorghum (Sorghum biocolor) as an energy crop with the focus on the biomass or non-grain components of the plant. The project will conduct basic studies on the genetic, biochemical, and physiological mechanisms affecting the composition of sorghum biomass and its conversion to liquid fuels. The focus will be on decreasing or increasing lignin content and/or modifying its composition and on increasing sugar content and yield for juice extraction. Low lignin is desirable for the saccharification and fermentation conversion process while high lignin concentration is desirable for conversion via pyrolysis. The impacts of fungal pathogens on sorghum with compositionally modified biomass will be determined. Germplasm with desirable genes affecting the conversion of sorghum biomass to energy will be developed, fully characterized, and released and deposited in Germplasm Resources Information Network (GRIN) for use by public and private sector plant breeders for developing improved hybrids and cultivars. The project consists of three integrated components: germplasm development, molecular biology, and plant pathology (Figure 1). Molecular and conventional methodologies will be utilized, and the project scale will range from field-level to gene-level. The project also has extensive formal and informal collaborations enhancing our ability to conduct this research. Anticipated products include improved sorghum germplasm for the sorghum seed industry with enhanced energy traits and biotic stress tolerance, and tools to assess the biological pathways that impact bioenergy traits and fungal pathogen responses of sorghum.

3. Progress Report:
This five-year project ended February of 2018, and was renewed as project number 3042-21220-033-00D, Genetic Improvement of Sorghum for Bioenergy, Feed, and Food Uses. During the course of this project, the following key accomplishments were achieved over the past 5 years and are summarized here. Lignin, a key component of plant cell walls that affects the efficiency of bioenergy conversion. In objective 1, structure and function of seven key enzymes required for lignin synthesis in sorghum were determined. Six novel bmr mutants were discovered, which reduce lignin content and increase breakdown cell walls into sugars. These mutants provide scientists with new tools to improve sorghum biomass for bioenergy and forage uses, and will potentially aid the identification of genes controlling lignin synthesis in sorghum and other bioenergy grasses. A Myb transcription factor, SbMyb60 was discovered that activates lignin synthesis in sorghum. Overexpression of SbMyb60 activated lignin biosynthesis pathway and other biochemical pathways required for producing lignin. SbMyb60 represents a tool to increase lignin and its intermediates in the cell walls of sorghum and other bioenergy grasses. Sorghum developed in this objective have served as the basis for a full range of scientific research from basic cell wall biochemistry to new forage and bioenergy applications, which will allow scientists to develop new strategies to modify lignin content and its composition in sorghum and other grasses. Stalk pathogens can be particularly devastating to sorghum production, and identifying sources of resistance is challenging because numerous types of fungi are responsible for stalk rot diseases. In Objective 2, whether bmr6 or bmr12 affected the susceptibility of sorghum to stalk diseases, Fusarium stalk rot and charcoal rot were evaluated, because lignin accumulation is a common response to pathogens. These studies examined plant background, pathogens, phenolic compounds, stalk extracts, and bmr mutations, which together provide a complete picture of stalk rot resistance. This body of research showed that bmr6 and bmr12 do not increase susceptibility to stalk diseases, but instead bmr6 and bmr12 have increased stalk rot resistance under some conditions. bmr plants provide a novel basis for scientists to develop new strategies to combat stalk rot. Sweet sorghum varieties were examined for resistance to stalk rot pathogen, which led to the identification of resistant lines. In sorghum grain, Waxy, the sorghum granule-bound starch synthase produces amylose starch. waxy lines are valuable for enhancing sorghum grain for bioenergy conversion and food processing. In a series of studies, susceptibility waxy and normal sorghum to grain mold fungi were examined, and similar studies were also performed with waxy wheat. The results for both crops showed that waxy lines were not more susceptible to grain infections than normal lines. This research demonstrated that these traits to enhance bioenergy availability for grain, biomass and stalk sugars do not affect sorghum resistance to these fungal pathogens, which allows breeders to develop resistant sorghums to meet a range of end-uses. In Objective 3, sorghum lines containing the waxy mutations were developed and released for the production sorghum grain with modified starch. Waxy hybrids were also developed using these lines, which are some of the first publically-available germplasm that can produce high-yielding waxy hybrids. These lines were shown to have greater digestibility of the grain for livestock feed and conversion efficiency to ethanol. These hybrids are being commercially produced and evaluated by U.S. food companies. The waxy trait opens sorghum grain to new uses, markets and applications for U.S. producers. In grazing trial, bmr12 was shown to increased beef cattle performance compared to those cattle fed on normal sorghum stalks. This research demonstrates the increased digestibility associated with bmr12 translated to increased cattle performance in the field. Germplasm released from the project is impacting both basic science and the sorghum industry.

4. Accomplishments
1. Discovery of a sorghum transcription factor that controls lignin synthesis. Lignin is the major structural component of plant cell walls whose presence and composition influences the usability of plant biomass for the production of biofuels and other natural products. ARS scientists at Lincoln, Nebraska discovered a Myb transcription factor, SbMyb60 that activates synthesis of lignin and other biochemical pathways in sorghum. Experiments showed that overexpression of SbMyb60 increased the synthesis of the amino acid phenylalanine and other compounds required for lignin synthesis, activated genes in the lignin biosynthesis pathway, and led to increased lignin levels in sorghum biomass. SbMyb60 is the first activator of lignin synthesis to be identified in grasses. SbMyb60 was discovered to not only affect lignin synthesis, but also redirect plant metabolism towards its production. SbMyb60 represents a tool to modify plant cell wall composition and the potential to improve biomass for renewable uses in sorghum and other bioenergy grasses.

2. Reducing lignin of sorghum does not increase its susceptibility to stalk rot or grain mold. Reducing lignin increases the conversion efficiency of biomass into sugars, but lignin is important for plant defenses against pathogens. Stalk rot fungi reduce grain and biomass yields, and the destructiveness of these diseases increases under drought conditions. ARS scientists in Lincoln, Nebraska investigated whether brown midrib (bmr) mutants, which have reduced lignin levels, change the susceptibility of sorghum to fungal diseases in an extensive series of studies. Several bmr and normal sorghum lines were tested for their susceptibility to the fungal pathogens that cause stalk rot diseases and grain molds. None of the bmr lines showed increased susceptibility to these pathogens, and in fact some bmr lines were more resistant compared to their normal counterparts. In addition, the increased resistance observed is driving research toward the identification of biochemical compounds or other factors that inhibit these fungal diseases. Susceptibility of bmr lines to stalk rot pathogens under drought conditions has examined where the disease can be particularly devastating, and again some bmr lines show increased under these conditions. The source of increased resistance in some bmr lines is under investigation as a potential crop protection tool. These studies demonstrated that bmr lines do not increase susceptibility of sorghum to diseases, which reduce yield and negatively impact biomass/forage quality for end-users. Therefore, bmr lines can be used to enhance sorghum for bioenergy and livestock feed.

3. The development and release of waxy sorghum lines for the food and ethanol industries. Starch is normally composed of two types of molecules, amylose and amylopectin. Waxy starch is almost entirely composed of amylopectin, which changes the physical properties of the starch making it more useful for the food and ethanol industries. ARS scientists at Lincoln, Nebraska developed, evaluated, and released a series of waxy sorghum lines. These lines were also combined to produce sorghum hybrids, and the hybrids and their parents were evaluated for grain yield, starch composition and susceptibility to grain molds. The waxy grain is also more digestible than normal grain, so there was a concern that waxy grain would be more susceptible to grain molds. These studies demonstrated that waxy sorghum grain is not more susceptible to grain mold, and the waxy hybrids can produce yields comparable to normal hybrids. These lines are some of the only publically-available germplasm capable of producing high-yielding waxy hybrids. The release of this waxy germplasm has enabled the commercial production waxy sorghum and open sorghum grain to new industrial food applications.

Review Publications
Funnell-Harris, D.L., Scully, E.D., Sattler, S.E., French, R.C., ONeill, P.M., Pedersen, J.F. 2017. Differences in fusarium species in brown midrib sorghum and in air populations in production fields. Phytopathology. 107(11):1353-1363.
Palmer, N.A., Saathoff, A.J., Scully, E.D., Tobias, C.M., Twigg, P., Madhavan, S., Schmer, M.R., Cahoon, R., Sattler, S.E., Edme, S.J., Mitchell, R., Sarath, G. 2017. Seasonal below-ground metabolism in switchgrass. Plant Journal. 92(6):1059-1075.
Scully, E.D., Gries, T.L., Palmer, N.A., Sarath, G., Funnell-Harris, D.L., Baird, L., Twigg, P., Seravalli, J., Clemente, T.E., Sattler, S.E. 2017. Overexpression of SbMyb60 in sorghum bicolor impacts both primary and secondary metabolism. New Phytologist. 217(1):82-104. doi:10.1111/nph.14815.