Lignin is an abundant biopolymer with a high carbon content and high aromaticity. Despite its potential as a raw material for the fuel and chemical industries, lignin remains the most poorly utilised of the lignocellulosic biopolymers. Effective valorisation of lignin requires careful fine-tuning of multiple "upstream" (i.e., lignin bioengineering, lignin isolation and "early-stage catalytic conversion of lignin") and "downstream" (i.e., lignin depolymerisation and upgrading) process stages, demanding input and understanding from a broad array of scientific disciplines. This review provides a "beginning-to-end" analysis of the recent advances reported in lignin valorisation. Particular emphasis is placed on the improved understanding of lignin's biosynthesis and structure, differences in structure and chemical bonding between native and technical lignins, emerging catalytic valorisation strategies, and the relationships between lignin structure and catalyst performance.

Paving the Way for Lignin Valorisation: Recent Advances in Bioengineering, Biorefining and Catalysis.

Trends in bioconversion of lignocellulose: Biofuels, platform chemicals & biorefinery concept

With the arising of global climate change and resource shortage, in recent years, increased attention has been paid to environmentally friendly materials. Trees are sustainable and renewable materials, which give us shelter and oxygen and remove carbon dioxide from the atmosphere. Trees are a primary resource that human society depends upon every day, for example, homes, heating, furniture, and aircraft. Wood from trees gives us paper, cardboard, and medical supplies, thus impacting our homes, school, work, and play. All of the above-mentioned applications have been well developed over the past thousands of years. However, trees and wood have much more to offer us as advanced materials, impacting emerging high-tech fields, such as bioengineering, flexible electronics, and clean energy. Wood naturally has a hierarchical structure, composed of well-oriented microfibers and tracheids for water, ion, and oxygen transportation during metabolism. At higher magnification, the walls of fiber cells have an interes...

https://www.fpl.fs.fed.us/documnts/pdf2016/fpl_2016_zhu001.pdf

Wood-Derived Materials for Green Electronics, Biological Devices, and Energy Applications.

Soil Nutrient Bioavailability—A Mechanistic Approach

Background
Agricultural production is often limited by low phosphorus (P) availability. In developing countries, which have limited access to P fertiliser, there is a need to develop plants that are more efficient at low soil P. In fertilised and intensive systems, P-efficient plants are required to minimise inefficient use of P-inputs and to reduce potential for loss of P to the environment.

Plant and microbial strategies to improve the phosphorus efficiency of agriculture

Switchgrass is a leading dedicated bioenergy feedstock in the United States because it is a native, high-yielding, perennial prairie grass with a broad cultivation range and low agronomic input requirements Biomass conversion research has developed processes for production of ethanol and other biofuels, but they remain costly primarily because of the intrinsic recalcitrance of biomass We show here that genetic modification of switchgrass can produce phenotypically normal plants that have reduced thermal-chemical (≤180 °C), enzymatic, and microbial recalcitrance Down-regulation of the switchgrass caffeic acid O-methyltransferase gene decreases lignin content modestly, reduces the syringyl:guaiacyl lignin monomer ratio, improves forage quality, and, most importantly, increases the ethanol yield by up to 38% using conventional biomass fermentation processes The down-regulated lines require less severe pretreatment and 300–400% lower cellulase dosages for equivalent product yields using simultaneous saccharification and fermentation with yeast Furthermore, fermentation of diluted acid-pretreated transgenic switchgrass using Clostridium thermocellum with no added enzymes showed better product yields than obtained with unmodified switchgrass Therefore, this apparent reduction in the recalcitrance of transgenic switchgrass has the potential to lower processing costs for biomass fermentation-derived fuels and chemicals significantly Alternatively, such modified transgenic switchgrass lines should yield significantly more fermentation chemicals per hectare under identical process conditions

https://www.pnas.org/content/pnas/108/9/3803.full.pdf

Genetic manipulation of lignin reduces recalcitrance and improves ethanol production from switchgrass

The bioconversion of carbohydrates in the herbaceous bioenergy crop, switchgrass (Panicum virgatum L.), is limited by the associated lignins in the biomass. The cinnamyl alcohol dehydrogenase (CAD) gene encodes a key enzyme which catalyzes the last step of lignin monomer biosynthesis. Transgenic switchgrass plants were produced with a CAD RNAi gene construct under the control of the maize ubiquitin promoter. The transgenic lines showed reduced CAD expression levels, reduced enzyme activities, reduced lignin content, and altered lignin composition. The modification of lignin biosynthesis resulted in improved sugar release and forage digestibility. Significant increases of saccharification efficiency were obtained in most of the transgenic lines with or without acid pretreatment. A negative correlation between lignin content and sugar release was found among these transgenic switchgrass lines. The transgenic materials have the potential to allow for improved efficiency of cellulosic ethanol production.

Downregulation of Cinnamyl Alcohol Dehydrogenase (CAD) Leads to Improved Saccharification Efficiency in Switchgrass

Improving phosphorus acquisition of white clover (Trifolium repens L.) by transgenic expression of plant-derived phytase and acid phosphatase genes.

Switchgrass (Panicum virgatum L.) has been developed into an important biofuel crop. Embryogenic calli induced from caryopses or inflorescences of the lowland switchgrass cultivar Alamo were used for Agrobacterium-mediated transformation. A chimeric hygromycin phosphotransferase gene (hph) was used as the selectable marker and hygromycin as the selection agent. Embryogenic calli were infected with Agrobacterium tumefaciens strain EHA105. Calli resistant to hygromycin were obtained after 5 to 8 weeks of selection. Soil-grown transgenic switchgrass plants were obtained 4 to 5 months after Agrobacterium infection. The transgenic nature of the regenerated plants was demonstrated by PCR, Southern blot hybridization analysis, and GUS staining. T1 progeny were obtained after reciprocal crosses between transgenic and untransformed control plants. Molecular analyses of the T1 progeny revealed various patterns of segregation. Transgene silencing was observed in the progeny with multiple inserts. Interestingly, reversal of the expression of the silenced transgene was found in segregating progeny with a single insert.

Agrobacterium -Mediated Transformation of Switchgrass and Inheritance of the Transgenes

The development of genome editing technologies offers new prospects in improving bioenergy crops like switchgrass (Panicum virgatum). Switchgrass is an outcrossing species with an allotetraploid genome (2n = 4x = 36), a complexity which forms an impediment to generating homozygous knock-out plants. Lignin, a major component of the plant cell wall and a contributor to cellulosic feedstock’s recalcitrance to decomposition, stands as a barrier to efficient biofuel production by limiting enzyme access to cell wall polymers during the fermentation process. We developed a CRISPR/Cas9 genome editing system in switchgrass to target a key enzyme involved in the early steps of monolignol biosynthesis, 4-Coumarate:coenzyme A ligase (4CL). Three 4CL genes, Pv4CL1, Pv4CL2, and Pv4CL3, were identified in switchgrass. Expression analysis revealed that Pv4CL1 transcripts were more abundant in the stem than in the leaf, while Pv4CL2 transcripts were barely detectable and Pv4CL3 was mainly expressed in the leaf. Pv4CL1 was selected as the target for CRISPR/Cas9 editing because of its preferential expression in highly lignified stem tissues. Specific guide RNA was constructed to target Pv4CL1. After introducing the construct into switchgrass calli, 39 transgenic plants were regenerated. Using two rounds of PCR screening and sequencing, four plants were confirmed to have tetra-allelic mutations simultaneously. The Pv4CL1 knock-out plants had reduced cell wall thickness, an 8–30% reduction in total lignin content, a 7–11% increase in glucose release, and a 23–32% increase in xylose release. This study established a successful CRISPR/Cas9 system in switchgrass with mutation efficiency reaching 10%. The system allows the precise targeting of the selected Pv4CL1 gene to create switchgrass knock-out mutant plants with decreased lignin content and reduced recalcitrance.

/pdf/defined-tetra-allelic-gene-disruption-of-the-4-coumarate-3rxlwxfjmh.pdf

Defined tetra-allelic gene disruption of the 4-coumarate:coenzyme A ligase 1 (Pv4CL1) gene by CRISPR/Cas9 in switchgrass results in lignin reduction and improved sugar release

Yaxin Ge

Papers

Genetic manipulation of lignin reduces recalcitrance and improves ethanol production from switchgrass

Downregulation of Cinnamyl Alcohol Dehydrogenase (CAD) Leads to Improved Saccharification Efficiency in Switchgrass

Improving phosphorus acquisition of white clover (Trifolium repens L.) by transgenic expression of plant-derived phytase and acid phosphatase genes.

Agrobacterium -Mediated Transformation of Switchgrass and Inheritance of the Transgenes

Defined tetra-allelic gene disruption of the 4-coumarate:coenzyme A ligase 1 (Pv4CL1) gene by CRISPR/Cas9 in switchgrass results in lignin reduction and improved sugar release