Deficit irrigation is increasingly used to reduce agricultural water use in arid regions, yet reliable field-scale quantification of consumptive water savings under commercial conditions remains challenging. Most satellite-based evapotranspiration (ET) studies focus on well-watered systems, whereas deficit irrigation imposes distinct soil-canopy controls on ET response. This study evaluated OpenET satellite-derived actual crop ET (ETc act) for quantifying summer deficit-irrigation impacts in alfalfa systems of California's Imperial Valley using a two-year field analysis (2024-2025) across 20 grower-managed fields. OpenET ETc act estimates were benchmarked under full irrigation using eddy covariance and under deficit irrigation using SM-RZD. Under full irrigation, OpenET products reproduced daily ETc act dynamics with good agreement, with mean bias errors generally below 0.5 mm d(-)& sup1; . Following irrigation cutoff, ETc act declined from 6 to 9 mm d(-)& sup1; to sustained levels near 1-3 mm d(-)& sup1; , driven primarily by soil-water depletion and reduced transpiration rather than atmospheric demand, supported by declines in NDVI and K-c act. Deficit irrigation reduced cumulative ETc act by 150-200 mm per deficit period, corresponding to 40-50% lower consumptive water use relative to full irrigation (p < 0.001). Despite rainfall variability during the wetter season, deficit-irrigated fields exhibited significantly lower cumulative ET, and the OpenET ensemble provided the most stable representation across conditions. These results indicate that satellite-derived ETc act, when evaluated under water-limited conditions and interpreted using soil-moisture and canopy indicators, provides a credible and operational basis for verifying consumptive water savings from deficit irrigation in arid agricultural systems.

Conversion of lignocellulosic biomass to renewable biofuels and bio-based chemicals at scale holds great promise for increasing domestic supplies of energy and alternatives to petrochemicals but is contingent upon improvements in biomass deconstruction. Here we report a method to maximize the enzymatic hydrolysis of whole plant samples containing fibrous and non-fibrous carbohydrates in sorghum, a naturally drought-tolerant high-yielding bioenergy feedstock. This process involves the addition of amylases to ionic liquid-pretreated biomass in conjunction with traditionally employed cellulase and hemicellulase cocktails. We found that amylases are compatible with cholinium-based ionic liquids and act synergistically with other saccharolytic enzymes to deconstruct sorghum biomass. By using mixtures of sorghum grain (high starch content) and stover (low starch content), we show that amylases can produce up to 4-fold higher glucose yields from biomass with a higher starch content but even samples containing only stover can benefit from amylase addition. Total monomeric sugar concentrations of 70 and 153 g/L were obtained from stover or grain biomass, respectively, when performing a reaction with 20

Winter groundwater recharge, which involves flooding farmland with excess surface water during the dormant season to replenish underlying aquifers, is a promising water conservation strategy, and alfalfa represents a particularly promising crop for this practice. In this study, we investigated yield and forage quality effects of intentional winter or spring flooding of non-dormant alfalfa (Medicago sativa L.) for groundwater recharge. A replicated randomized complete block design study was implemented at the University of California Kearney Research and Extension Center (KARE) in Parlier, CA testing three treatments (3 days of flooding followed by 4 days without flooding, 4 days of flooding followed by 10 days without flooding, and control) for a duration of 6 weeks in winter 2019 and spring 2020. A total of 1.6 and 3.6 m(3) m(-2) (2019) and 1.3 and 2.2 m(3) m(-2) (2020) were recharged in the 4 on 10 off and 3 on 4 off flood treatments at a recharge efficiency of > 88 % (81 % - 217 % of annual irrigation demand), respectively. The 2019 (Feb - Mar) experiment did not show significant differences in alfalfa yield in the first and second cutting after flooding but resulted in slight but non-significant decline in forage quality (e.g. crude protein; fair hay quality). The 2020 flooding experiment, conducted much later in the year (April - May), resulted in no significant differences in alfalfa yield but poor feed quality (utility grade), partly due to delays in harvesting. Together these results indicate that while caution is appropriate to prevent oxygen deprivation and impacts on alfalfa yield and quality, winter recharge in alfalfa fields in highly permeable soils appears to be a viable practice to conserve large quantities of surface water when available in excess.

BackgroundThe development of alfalfa cultivars with improved digestibility may minimize the yield-quality tradeoff, enabling higher quality with late-harvested forage and possibly higher yields. MethodsAn irrigated experiment conducted over 4 years compared 28-d harvest schedules with 35-d harvest schedules and an alternating 21-d and 35-d schedule. Four conventional cultivars and four cultivars developed for higher digestibility were grown under each schedule. ResultsDelayed cutting (35-d) yields were 16% greater and the staggered treatments were 6% higher than the 28-d strategy. The nutritive value decreased significantly with the 35-d schedule, but a "staggered" system provided nutritive value similar to the 28-d schedule while achieving higher yields. The nutritive value of cultivars was in the order of HarvXtra>Hi-Gest> conventional cultivars. The HarvXtra but not Hi-Gest cultivars achieved similar digestibility under the 35-d cutting schedule compared with conventional cultivars on a 28-d schedule. ConclusionsThis study clearly demonstrates that higher nutritive value cultivars of fall dormancy 6-9 grown with staggered or late cutting schedules can increase yields while maintaining higher nutritive value. The combination of staggered or late schedules with improved cultivars can maximize yields while maintaining the nutritive value of alfalfa, potentially breaking the alfalfa yield-quality tradeoff.

Sorghum cultivars, particularly those used for forage and biomass, present significant potential as drought-resistant crops suitable for animal feed and biofuel production. This study evaluated 59 sorghum hybrids over five years (2019–2023) across three University of California research farm locations in the Central Valley: Kearney REC (KARE), West Side REC (WSREC), and Davis. The primary aim was to identify genotypes that exhibit high yield and stability across diverse environments in California, which is crucial for meeting the state’s significant feed needs associated with dairy operations and animal production. The evaluation focused on biomass yields, forage quality traits such as Relative Feed Quality (RFQ) and milk yield per ton (milk/ton), and biofuel-relevant chemical compositions like Neutral Detergent Fiber (NDF) and starch. A multi-trait stability index was employed to pinpoint superior genotypes that combine high yield with desirable quality traits. Results indicated significant genotypic, environmental, and genotype-by-environment (GxE) interaction effects for all traits except fat and water-soluble sugars. Eight hybrids were notable for maintaining high and stable biomass yields across different locations. Additionally, high fat and starch content were found to correlate with improved milk/ton potential, while lower fiber content (ADF, NDF) was associated with enhanced RFQ. Specifically, nine hybrids were identified as optimal for dairy forage due to their combination of high yield, RFQ, and milk/ton. Furthermore, distinct hybrids were identified for first-generation (starch-based) and second-generation (NDF-based) biofuel strategies. Three hybrids stood out as having desirable traits for both feed and biofuel applications, underscoring their versatility. This study highlights the utility of a multi-trait stability index in selecting superior sorghum genotypes for specific trait combinations. The identified candidates for forage and biofuel use, especially the multipurpose varieties, offer valuable insights that can aid growers and industry stakeholders in developing more sustainable and versatile sorghum production systems in California. Findings from this study contribute significantly to the development of more resilient sorghum production systems. By identifying hybrids that excel in both yield and quality across various environments, this research supports future cropping decisions aimed at enhancing water use efficiency and drought resilience in sorghum cultivation. These advancements are crucial for maintaining competitive dairy operations and advancing biofuel production in the face of climate change-induced challenges.

This study aims to detect different irrigation treatments in sorghum plant breeding experiments using multispectral satellite imagery coupled with a random forest classification model. Obtained from Planet Labs' official repository, satellite images span three sorghum test plots in Central Valley, CA, USA, covering a range of soil and climatological conditions. At each test plot, three irrigation treatments were applied: 100% full irrigation, 70% reduction, and 50% reduction to assess the drought resistance of different sorghum cultivars. A Random Forest Classification algorithm was used to detect the different treatments with high accuracy across all sites. The findings illustrate (1) the predictive capacity of PlanetScope Multispectral imagery for differences in irrigation across different sites and cultivars of sorghum, (2) the efficacy of Random Forest methods in accurately distinguishing diverse irrigation treatments using multispectral satellite imagery, and (3) the demonstrable advantage of utilizing multitemporal data over single-day imagery-based classification approaches. This work is an important step in advancing the utility of satellite remote sensing in irrigation monitoring water resources management, plant breeding studies, and supporting precision agriculture applications.

Abstract Background Lignin is an aromatic polymer deposited in secondary cell walls of higher plants to provide strength, rigidity, and hydrophobicity to vascular tissues. Due to its interconnections with cell wall polysaccharides, lignin plays important roles during plant growth and defense, but also has a negative impact on industrial processes aimed at obtaining monosaccharides from plant biomass. Engineering lignin offers a solution to this issue. For example, previous work showed that heterologous expression of a coliphage S-adenosylmethionine hydrolase (AdoMetase) was an effective approach to reduce lignin in the model plant Arabidopsis. The efficacy of this engineering strategy remains to be evaluated in bioenergy crops. Results We studied the impact of expressing AdoMetase on lignin synthesis in sorghum (Sorghum bicolor L. Moench). Lignin content, monomer composition, and size, as well as biomass saccharification efficiency were determined in transgenic sorghum lines. The transcriptome and metabolome were analyzed in stems at three developmental stages. Plant growth and biomass composition was further evaluated under field conditions. Results evidenced that lignin was reduced by 18% in the best transgenic line, presumably due to reduced activity of the S-adenosylmethionine-dependent O-methyltransferases involved in lignin synthesis. The modified sorghum features altered lignin monomer composition and increased lignin molecular weights. The degree of methylation of glucuronic acid on xylan was reduced. These changes enabled a ~20% increase in glucose yield after biomass pretreatment and saccharification compared to wild type. RNA-seq and untargeted metabolomic analyses evidenced some pleiotropic effects associated with AdoMetase expression. The transgenic sorghum showed developmental delay and reduced biomass yields at harvest, especially under field growing conditions. Conclusions The expression of AdoMetase represents an effective lignin engineering approach in sorghum. However, considering that this strategy potentially impacts multiple S-adenosylmethionine-dependent methyltransferases, adequate promoters for fine-tuning AdoMetase expression will be needed to mitigate yield penalty.

Sorghum is an attractive feedstock for biobased fuel and chemical production because it is familiar to farmers, naturally drought tolerant, and versatile as a food, feed, and fuel crop. Although sorghum is a promising feedstock, particularly in regions that experience drought stress, little is known about how drought conditions impact the ease of conversion of sorghum to fuels and products. This study combines agronomic field trials with a high-throughput experimental pipeline to explore the field performance and liquid biofuel (bisabolene) yields resulting from three sorghum types (photosensitive forage sorghum, optimized grain sorghum, and drought-resistant grain sorghum) grown under pre- and postflowering water limitations in two different California locations. Multiple drought treatments are compared to the control, as the timing (preflowering versus postflowering) of drought stress elicits different survival strategies and corresponding impacts on yield and composition. Forage-type sorghum maintained the highest biomass yields across all irrigation conditions and locations. Glucose and xylose yields resulting from ionic liquid pretreatment and enzymatic saccharification were not significantly impacted by irrigation treatments but differed by location and genotype. However, Rhodosporidium toruloides grown on the resulting plant hydrolysates unexpectedly produced higher titers of bisabolene for drought-stressed sorghum samples regardless of genotype.

A four-year research experiment was conducted on sandy loam soil at the University of California Kearney Agricultural Research and Extension Center in Parlier, California, to investigate the effect of midsummer deficit irrigation on alfalfa yield, irrigation water productivity (IWP), and crop water productivity (CWP). The experiment was a randomized block design with two treatments: full and deficit irrigations with three replications. Applied irrigation water was measured using flow meters and soil matric potentials were monitored using watermark soil moisture sensors. Actual evapotranspiration (ETa) values were estimated from Tule Technologies stations. The deficit irrigation treatments resulted in 454, 706, 625, and 815 mm of irrigation water savings as compared to the full irrigation treatments in 2019, 2020, 2021, and 2022, respectively. These values represent 30.3%, 40.9%, 37.0%, and 49.1% of the applied water savings. Alfalfa yield in the deficit treatments was reduced by 3.94, 2.04, 1.25, and 0.40 Mg ha-1; the equivalent of 18.1%, 11.1%, 7.1%, and 3.0% of the yield for the full irrigation treatment for the four years: with an average reduction of 10.7%. IWP was higher when deficit irrigation was implemented and resulted in 17.09, 16.11, 15.40, and 15.54 kg ha-1 mm-1, in 2019, 2020, 2021, and 2022, respectively. The production function using applied irrigation water (IW, mm) was: Y(yield in Mg ha-1)=0.50x(IW)2-1,633.75x(IW)+1,338,472 and Y=-0.1137x(IW)2+233.55x(IW)-103,036 for the full and deficit irrigation treatments, respectively. CWP was 18.6, 16.4, 14.9, and 12.3 kg ha-1 mm-1 for fully irrigated treatments, and 15.2, 14.9, 14.3, and 12.6 kg ha-1 mm-1 for the deficit irrigation treatments, for 2019, 2020, 2021, and 2022, respectively. Results from this work provide growers with viable deficit irrigation practices that could be implemented during drought periods.

Alfalfa is one of the major perennial forage crops in California, vital for the livestock industry, and provides environmental benefits to the ecosystem in the state. Information on subsurface drip irrigation (SDI) practices on alfalfa is limited particularly in areas related to drip tape spacings and installation depths and their impacts on topsoil profile wetting patterns and alfalfa productivity. The objective of this study was to compare three different depths of drip lines: 15, 30, and 45 cm on topsoil profile wetting patterns and identify the optimum depth for achieving sustainable management practices for alfalfa production under SDI. Crop water requirements were determined using Tule Technologies system. Volumetric soil water contents from twelve cuts were simulated using HYDRUS-2D. Initial volumetric soil water contents from Watermark sensors were used in the model and measured volumetric soil water distributions were used for hydraulic conductivity calibration. Simulated results showed that there was no significant difference between root water uptake (RWU) among the various drip depths. RWU was 171.6, 170.0, and 168.2 cm at drip line depths of 15, 30, and 45 cm, respectively. Applied irrigation water during the study period was 195.1 cm while rainfall was 4.1 cm. A 10% reduction in topsoil volumetric soil water content was observed for drip lines at 30 cm depth as compared with 15 cm depth, while a 20% reduction in topsoil volumetric soil water content was observed for drip lines at 45 cm depth as compared with 15 cm depth. Drip lines at 30 cm depth are likely the optimal for RWU and free drainage; however, drip lines at 45 cm depth may allow growers to provide additional irrigation events (without increasing the topsoil volumetric soil water content) closer to the harvest date, potentially resulting in higher yield and water use efficiency.

Background: Soil and water salinity are increasing problems worldwide, causing significantly reduced crop yields. Alfalfa (Medicago sativa L.) is often listed as salt-sensitive, but field testing of improved cultivars is limited. Forage systems and improved high-quality alfalfa varieties are needed to enable crop production under high salinity (HS) conditions. Methods: The objective of this study was to measure the yield and quality response of alfalfa to high saline conditions in the field and to document the relative saline tolerance of its varieties. HS irrigation water (electrical conductivity of water, or ECw 8.0-11.0dSm(-1)) was applied to 33 nondormant alfalfa cultivars and were compared with low salinity (LS) treatments (ECw 0.5-1.2dSm(-1)) over 4 years in a Mediterranean environment on a clay loam soil utilizing a split-plot design. Crops were harvested seven to eight times per year, and the forage quality was measured on selected harvests utilizing near-infrared spectroscopy. Results: The average yield loss due to HS treatment was 23.9% compared with LS treatment, but yields averaged 23.4Mgha(-1) under HS over the 3 full years of production. This level of production is considered to be economically viable in this region. Differences in salinity tolerance between lines were identified in the field; individual cultivars lost 5%-35% of their LS yield when grown under HS conditions. Forage quality was significantly improved under HS versus LS conditions, but improvements were negatively correlated with biomass yield (R-2>0.81), similar to responses observed in drought-stressed alfalfa. Conclusions: These yield results confirm greenhouse studies, indicating that alfalfa is highly salt tolerant once established in the field, with potential for further improvement with tolerant cultivars. Salinity tolerance should be chosen based on total biomass yield as well as on the salinity tolerance index (HS yield relative to LS yield). Agronomic practices to mitigate salinity and sodicity are critical, along with improved cultivars.

Building a stronger bioeconomy requires production capabilities that can be generated through microbial genetic engineering. Engineered microbes can be paired with engineered feedstocks and compatible deconstruction methods to improve production.

Switchgrass (Panicum virgatum L.) is a promising perennial bioenergy crop that achieves high yields with relatively low nutrient and energy inputs. Modification of cell wall composition for reduced recalcitrance can lower the costs of deconstructing biomass to fermentable sugars and other intermediates. We have engineered overexpression of OsAT10, encoding a rice BAHD acyltransferase and QsuB, encoding dehydroshikimate dehydratase from Corynebacterium glutamicum, to enhance saccharification efficiency in switchgrass. These engineering strategies demonstrated low lignin content, low ferulic acid esters, and increased saccharification yield during greenhouse studies in switchgrass and other plant species. In this work, transgenic switchgrass plants overexpressing either OsAT10 or QsuB were tested in the field in Davis, California, USA for three growing seasons. No significant differences in the content of lignin and cell wall-bound p-coumaric acid or ferulic acid were detected in transgenic OsAT10 lines compared with the untransformed Alamo control variety. However, the transgenic overexpressing QsuB lines had increased biomass yield and slightly increased biomass saccharification properties compared to the control plants. This work demonstrates good performance of engineered plants in the field, and also shows that the cell wall changes in the greenhouse were not replicated in the field, emphasizing the need to validate engineered plants under relevant field conditions.

Glyphosate-resistant (GR) alfalfa (Medicago sativa L.) has been widely adopted in the United States. Exceptional tolerance of GR alfalfa to glyphosate application has been reported as a strength of this technology; however, growers have recently reported potential crop injury under specific environmental conditions. The purpose of this study was to document and characterize the injury and determine best management practices for avoiding injury to GR alfalfa in the Intermountain West. The effects of glyphosate rate and application timing during various seasons were investigated at 24 sites over 5 years, measuring the impact on alfalfa crop height and biomass yield. Summer glyphosate applications did not injure alfalfa. However, spring applications reduced crop height at 76% of sites and biomass yield at 62% of sites. At responsive sites, low (869 g ae ha(-1)) and high (1739 g ae ha(-1)) glyphosate rates reduced yield by 0.53 and 1.06 Mg ha(-1), respectively. Alfalfa treated with high rates in the late spring, when 15-20 cm tall, had mean yield reductions of 16%-17% compared with untreated alfalfa. These results suggest that glyphosate applications made at tall crop heights or high rates on GR alfalfa are more likely to reduce crop height and yield in the Intermountain West compared with earlier applications at lower rates. We recommend that spring applications using low glyphosate rates occur before alfalfa is 10 cm tall to mitigate the risk of injury. If a high glyphosate rate is necessary, then an application should be made before alfalfa is greater than 5 cm tall.

Abstract Postharvest blanching and drying of industrial hemp (Cannabis sativa L.) by infrared (IR) and hot air (HA) heating was studied. Experiments were conducted at different IR heating times (1 and 2 min) and HA temperatures (65 and 85 °C) and compared with conventional indoor drying. Drying time was decreased from 3366 min (conventional) to as low as 222 min (85 °C HA). IR and HA processing reduced the total aerobic bacteria, and total yeast/mold levels by up to 0.81 and 1.85 log CFU/g, respectively from their initial levels of 4.63 and 4.75 log CFU/g, meanwhile reduced the activities of polyphenol oxidase and peroxidase by up to 91.7% and 66.7%, respectively. More than 96.1% total cannabidiol was preserved by thermal processing. Total terpene retention ranged from 18.3% to 71.1% under tested conditions with distinct terpene profiles. The results provide important information on the microbial safety and a timely solution to improve the postharvest processes of hemp.

Background Alfalfa is a globally important forage crop. Cultivars are characterized by fall dormancy (FD). FD affects biomass yield and winter survival and is used to identify appropriate growing regions of cultivars. It has historically been assessed by measuring the natural height of regrowth in autumn of spaced plants in the field. Because commercial alfalfa is normally grown as a solid planted sward, FD could be different if plants were grown under real production conditions. The objective of this experiment was to assess whether FD ratings obtained from swards were similar to those obtained on spaced plants. Methods We evaluated 20 alfalfa cultivars in field trials established in 2015 at five locations in the United States. We harvested the trials in early autumn and measured regrowth plant height approximately 3 weeks later in 2015 and 2016. Results Autumn plant height responded as expected for the established check cultivars in all locations. Fall dormancy ratings for cultivars under both management systems were highly correlated. Conclusions Estimating FD from spaced plant height measurements in autumn is very robust, and using height data from sward plots gives equivalent results to that measured in spaced plant nurseries. This finding has many practical advantages.

Agricultural Managed Aquifer Recharge (Ag-MAR) is a potential and sustainable practice where agricultural fields can be used to recharge depleted aquifers using excess precipitation during winter. However, there is little information on the amount of Ag-MAR that can be applied to crops such as alfalfa. HYDRUS-2D was used to estimate the net recharge in an alfalfa field grown on a sandy loam soil in a Mediterranean climate at Parlier, California, USA in 2020-2022. The alfalfa field had four irrigation treatments: full irrigation during summer growing season (March through November), mid-summer deficit irrigation treatment (March to August and complete irrigation cutoff after August cutting), winter flooding treatment, and no winter flooding. Recharge, evapotranspiration (ETa), soil moisture dynamics, and root water uptake were simulated during the recharge period in winter. Previously fully irrigated treatments in summer, followed by winter recharge led to cumulative groundwater recharge of 1459, 1687, and 1415 mm for 2020, 2021, and 2022, respectively. These applications resulted in a net recharge of 85%, 89%, and 84% of the applied irrigation water during the winter period, a significant contribution to groundwater aquifers. Mid-summer deficit irrigation treatments, followed by winter recharge, resulted in net groundwater recharge of 1337, 1498, and 1272 mm for 2020, 2021, and 2022, respectively, amounting to 78%, 79%, and 76% of the applied irrigation water during winter flooding periods. HYDRUS simulation model predicted groundwater recharge potential in these experiments successfully with a coefficient of determination, R2 values of 0.91, and 0.89 for the groundwater recharge during winter flooding after the full irrigation in summer, and the mid-summer deficit irrigation, respectively. These results confirm the potential utilization of HYDRUS simulations in predicting groundwater recharge potential under similar sandy -soil conditions in California's San Joaquin Valley.

Drought and climate change have decreased water availability for agriculture, especially in the desert of southwestern USA. Efficiency enhancements in irrigation management aimed at conserving water are key to adjust to limits in water supply, improve profitability and sustainability of alfalfa production in arid and semiarid areas. This study intended to conduct a field-scale analysis to develop yield and ET estimation tools for the effective use of irrigation water in a desert alfalfa production system. Extensive data collection and trials were carried out over three years in nine fully irrigated commercial alfalfa fields in the low deserts of California. The seasonal crop water consumption measured using the residual of energy balance method varied from 1381 mm to 1596 mm across the experimental sites and crop seasons. Variable seasonal dry mater (DM) yields ranged from 23.01 Mg ha−1 to 29.90 Mg ha−1. The results indicated that the first five cuttings each year were the most productive cuttings with a mean DM value ranged between 3.29 (cut 1) and 4.21 (cut 4) Mg ha−1 but declined in later cuttings. An average annual water productivity (WP) value of 17.0 kg ha−1 mm−1 was determined across the sites varying from 15.5 to 18.9 kg ha−1 mm−1. The findings suggested that one may lose up to 1.44 Mg ha−1 alfalfa yields with moderate summer deficit irrigation strategies, using 40% less water applied than full irrigation practices over the summer period of July–September. A more severe summer water deficit, with no irrigation event over the summer period of July–September may result in a potential water savings of 0.234–0.246 (ha·m) ha−1 and 19–21% seasonal yield losses in the desert environment. This study describes the seasonal yield pattern, the crop water use-production function, and the crop coefficient values for various harvest cycles over the crop season. These tools may assist farmers to quantify water savings and estimate yield losses for more accurate and effective irrigation management strategies to meet water conservation objectives and for the resiliency of alfalfa production in the desert region.

Engineering bioenergy crops to accumulate coproducts in planta can increase the value of lignocellulosic biomass and enable a sustainable bioeconomy. In this study, we engineered sorghum with a bacterial gene encoding a chorismate pyruvate-lyase (ubiC) to reroute the plastidial pool of chorismate from the shikimate pathway into the valuable compound 4-hydroxybenzoic acid (4-HBA). A gene encoding a feedback-resistant version of 3-deoxy-d-arabino-heptulonate-7-phosphate synthase (aroG) was also introduced in an attempt to increase the carbon flux through the shikimate pathway. At the full maturity and senesced stage, two independent lines that co-express ubiC and aroG produced 1.5 and 1.7 dw% of 4-HBA in biomass, which represents 36- and 40-fold increases compared to the titer measured in wildtype. The two transgenic lines showed no obvious phenotypes, growth defects, nor alteration of cell wall polysaccharide content when cultivated under controlled conditions. In the field, when harvested before grain maturity, transgenic lines contained 0.8 and 1.2 dw% of 4-HBA, which represent economically relevant titers based on recent technoeconomic analysis. Only a slight reduction (11-15%) in biomass yield was observed in transgenics grown under natural environment. This work provides the first metabolic engineering steps toward 4-HBA overproduction in the bioenergy crop sorghum to improve the economics of biorefineries by accumulating a value-added coproduct that can be recovered from biomass and provide an additional revenue stream.

Engineering bioenergy crops to accumulate value-added coproducts in planta is an attractive approach to increasing the value of lignocellulosic biomass and enabling a sustainable bioeconomy. In this study, we engineered sorghum with a bacterial gene encoding a dehydroshikimate dehydratase (qsuB) to convert the endogenous pool of 3-dehydroshikimate into the valuable compound protocatechuate (DHBA). We find that, when grown under field conditions, transgenic sorghum lines can accumulate up to 0.3% DHBA in stover on a dry weight (DW) basis without showing any difference in cell wall composition. An unexpected finding was an increase in yield for all qsuB-expressing lines. The grain yield and total biomass yield were 71 and 29% higher in the highest yielding line, respectively. On average, the total biomass yield of the engineered lines was 22.3 t/ha (DW). Moreover, we conducted a techno-economic analysis to investigate the economic impact of coproducing DHBA along with bioethanol in an integrated cellulosic biorefinery. Using engineered biomass sorghum with 0.3 DW% DHBA accumulated in planta as the feedstock, the economics of the integrated biorefineries has the potential to be improved. Our data demonstrate an engineering strategy to overproduce DHBA in bioenergy crops to facilitate sustainable manufacturing of biofuels and bioproducts.