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Gene stacking of multiple traits for high yield of fermentable sugars in plant biomass.

ABSTRACT: Background:Second-generation biofuels produced from biomass can help to decrease dependency on fossil fuels, bringing about many economic and environmental benefits. To make biomass more suitable for biorefinery use, we need a better understanding of plant cell wall biosynthesis. Increasing the ratio of C6 to C5 sugars in the cell wall and decreasing the lignin content are two important targets in engineering of plants that are more suitable for downstream processing for second-generation biofuel production. Results:We have studied the basic mechanisms of cell wall biosynthesis and identified genes involved in biosynthesis of pectic galactan, including the GALS1 galactan synthase and the UDP-galactose/UDP-rhamnose transporter URGT1. We have engineered plants with a more suitable biomass composition by applying these findings, in conjunction with synthetic biology and gene stacking tools. Plants were engineered to have up to fourfold more pectic galactan in stems by overexpressing GALS1, URGT1, and UGE2, a UDP-glucose epimerase. Furthermore, the increased galactan trait was engineered into plants that were already engineered to have low xylan content by restricting xylan biosynthesis to vessels where this polysaccharide is essential. Finally, the high galactan and low xylan traits were stacked with the low lignin trait obtained by expressing the QsuB gene encoding dehydroshikimate dehydratase in lignifying cells. Conclusion:The results show that approaches to increasing C6 sugar content, decreasing xylan, and reducing lignin content can be combined in an additive manner. Thus, the engineered lines obtained by this trait-stacking approach have substantially improved properties from the perspective of biofuel production, and they do not show any obvious negative growth effects. The approach used in this study can be readily transferred to bioenergy crop plants.

SUBMITTER: Aznar A

PROVIDER: S-EPMC5759196 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature

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Gene stacking of multiple traits for high yield of fermentable sugars in plant biomass.

Aznar Aude A Chalvin Camille C Shih Patrick M PM Maimann Michael M Ebert Berit B Birdseye Devon S DS Loqué Dominique D Scheller Henrik V HV

Biotechnology for biofuels 20180109

<h4>Background</h4>Second-generation biofuels produced from biomass can help to decrease dependency on fossil fuels, bringing about many economic and environmental benefits. To make biomass more suitable for biorefinery use, we need a better understanding of plant cell wall biosynthesis. Increasing the ratio of C6 to C5 sugars in the cell wall and decreasing the lignin content are two important targets in engineering of plants that are more suitable for downstream processing for second-generatio ...[more]

PMID: 29321811

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Project description:BackgroundPretreatment of biomass to maximize the recovery of fermentable sugars as well as to minimize the amount of enzyme inhibitors formed during the pretreatment is a challenge in biofuel process. We develop a modified Fenton pretreatment in a mixed solvent (water/DMSO) to combine the advantages of organosolv and Fenton pretreatments. The hemicellulose and cellulose in corncob were effectively degraded into xylose, glucose, and soluble glucose oligomers in a few hours. This saccharide solution, separated from the solid lignin simply by filtration, can be directly applied to the subsequent enzymatic hydrolysis and ethanol fermentation.ResultsAfter the pretreatment, 94% carbohydrates were recovered as soluble monosaccharide (xylose and glucose) and glucose oligomers in the filtrates, and 87% of solid lignin was recovered as the filter residue. The filtrates were directly applied to enzymatic hydrolysis, and 92% of raw corncob glucose was recovered. The hydrolysates containing the glucose and xylose from the enzymatic hydrolysis were directly applied to ethanol fermentation with ethanol yield equals 79% of theoretical yield. The pretreatment conditions (130 °C, 1.5 bar; 30 min to 4 h) are mild, and the pretreatment reagents (H2O2, FeCl3, and solvent) had low impact to environment. Using ferrimagnetic Fe3O4 resulted in similar pretreatment efficiency and Fe3O4 could be removed by filtration.ConclusionsA modified Fenton pretreatment of corncob in DMSO/water was developed. Up to 94% of the carbohydrate content of corncob was recovered as a saccharide solution simply by filtration. Such filtrate was directly applied to the subsequent enzymatic hydrolysis and where 92% of the corncob glucose content was obtained. The hydrolysate so obtained was directly applied to ethanol fermentation with good fermentability. The pretreatment method is simple, and the additives and solvents used have a low impact to the environment. This method provides the opportunity to substantially maximize the carbohydrate and solid lignin recovery of biomass with a comparatively green process, such that the efficiency of biorefinery as well as the bioethanol production process can be improved. The pretreatment is still relatively energy intensive and expensive, and further optimization of the process is required in large-scale operation.

Project description:A panel of 60 genotypes comprising New Plant Types (NPTs) along with indica, tropical and temperate japonica genotypes was phenotypically evaluated for four seasons in irrigated situation for grain yield per se and component traits. Twenty NPT genotypes were found promising with an average grain yield varying from 5.45 to 8.8 t/ha. A total of 85 SSR markers were used in the study to identify QTLs associated with grain yield per se and related traits. Sixty-six (77.65%) markers were found to be polymorphic. The PIC values varied from 0.516 to 0.92 with an average of 0.704. A moderate level of genetic diversity (0.39) was detected among genotypes. Variation to the tune of 8% within genotypes, 68% among the genotypes within the population and 24% among the populations were observed (AMOVA). This information may help in identification of potential parents for development of transgressive segregants with very high yield. The association analysis using GLM and MLM models led to the identification of 30 and 10 SSR markers associated with 70 and 16 QTLs, respectively. Thirty novel QTLs linked with 16 SSRs were identified to be associated with eleven traits, namely tiller number (qTL-6.1, qTL-11.1, qTL-4.1), panicle length (qPL-1.1, qPL-5.1, qPL-7.1, qPL-8.1), flag leaf length (qFLL-8.1, qFLL-9.1), flag leaf width (qFLW-6.2, qFLW-5.1, qFLW-8.1, qFLW-7.1), total no. of grains (qTG-2.2, qTG-a7.1), thousand-grain weight (qTGW-a1.1, qTGW-a9.2, qTGW-5.1, qTGW-8.1), fertile grains (qFG-7.1), seed length-breadth ratio (qSlb-3.1), plant height (qPHT-6.1, qPHT-9.1), days to 50% flowering (qFD-1.1) and grain yield per se (qYLD-5.1, qYLD-6.1a, qYLD-11.1).Some of the SSRs were co-localized with more than two traits. The highest co-localization was identified with RM5709 linked to nine traits, followed by RM297 with five traits. Similarly, RM5575, RM204, RM168, RM112, RM26499 and RM22899 were also recorded to be co-localized with more than one trait and could be rated as important for marker-assisted backcross breeding programs, for pyramiding of these QTLs for important yield traits, to produce new-generation rice for prospective increment in yield potentiality and breaking yield ceiling.

Project description:BackgroundLignocellulosic biomass has been commonly regarded as a potential feedstock for the production of biofuels and biochemicals. High sugar yields and the complete bioconversion of all the lignocellulosic sugars into valuable products are attractive for the utilization of lignocelluloses. It is essential to pretreat and hydrolyze lignocelluloses at high solids loadings during industrial processes, which is more economical and environmentally friendly as capital cost, energy consumption, and water usage can be reduced. However, oligosaccharides are inevitably released during the high solids loading enzymatic hydrolysis and they are more recalcitrant than monosaccharides for microorganisms.ResultsA fed-batch enzymatic hydrolysis of corn stover pretreated by the sodium hydroxide-methanol solution (SMs) at high solids loading was demonstrated to reach the high concentrations and yields of fermentable sugars. Glucose, xylose, cello-oligosaccharides, and xylo-oligosaccharides achieved 146.7 g/L, 58.7 g/L, 15.6 g/L, and 24.7 g/L, respectively, when the fed-batch hydrolysis was started at 12% (w/v) solids loading, and 7% fresh substrate and a standardized blend of cellulase, β-glucosidase, and hemicellulase were fed consecutively at 3, 6, 24, and 48 h to achieve a final solids loading of 40% (w/v). The total conversion of glucan and xylan reached 89.5% and 88.5%, respectively, when the oligosaccharides were taken into account. Then, a fed-batch culture on the hydrolysates was investigated for lipid production by Cutaneotrichosporon oleaginosum. Biomass, lipid content, and lipid yield were 50.7 g/L, 61.7%, and 0.18 g/g, respectively. The overall consumptions of cello-oligosaccharides and xylo-oligosaccharides reached 74.1% and 68.2%, respectively.ConclusionsHigh sugars concentrations and yields were achieved when the enzyme blend was supplemented simultaneously with the substrate at each time point of feeding during the fed-batch enzymatic hydrolysis. Oligosaccharides were co-utilized with monosaccharides during the fed-batch culture of C. oleaginosum. These results provide a promising strategy to hydrolyze alkaline organosolv-pretreated corn stover into fermentable sugars with high concentrations and yields for microbial lipid production.

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Gene stacking of multiple traits for high yield of fermentable sugars in plant biomass.

Publications

Gene stacking of multiple traits for high yield of fermentable sugars in plant biomass.

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