scholarly journals CRISPR-Knockout of CSE Gene Improves Saccharification Efficiency by Reducing Lignin Content in Hybrid Poplar

2021 ◽  
Vol 22 (18) ◽  
pp. 9750
Author(s):  
Hyun-A Jang ◽  
Eun-Kyung Bae ◽  
Min-Ha Kim ◽  
Su-Jin Park ◽  
Na-Young Choi ◽  
...  
Keyword(s):  
Lignocellulosic Biomass ◽  
Lignin Content ◽  
Hybrid Poplar ◽  
Lignin Biosynthesis ◽  
Biofuel Production ◽  
Guide Rnas ◽  
Hybrid Poplars ◽  
Four Seasons ◽  
Promising Target ◽  
Saccharification Efficiency

Caffeoyl shikimate esterase (CSE) has been shown to play an important role in lignin biosynthesis in plants and is, therefore, a promising target for generating improved lignocellulosic biomass crops for sustainable biofuel production. Populus spp. has two CSE genes (CSE1 and CSE2) and, thus, the hybrid poplar (Populus alba × P. glandulosa) investigated in this study has four CSE genes. Here, we present transgenic hybrid poplars with knockouts of each CSE gene achieved by CRISPR/Cas9. To knockout the CSE genes of the hybrid poplar, we designed three single guide RNAs (sg1–sg3), and produced three different transgenic poplars with either CSE1 (CSE1-sg2), CSE2 (CSE2-sg3), or both genes (CSE1/2-sg1) mutated. CSE1-sg2 and CSE2-sg3 poplars showed up to 29.1% reduction in lignin deposition with irregularly shaped xylem vessels. However, CSE1-sg2 and CSE2-sg3 poplars were morphologically indistinguishable from WT and showed no significant differences in growth in a long-term living modified organism (LMO) field-test covering four seasons. Gene expression analysis revealed that many lignin biosynthetic genes were downregulated in CSE1-sg2 and CSE2-sg3 poplars. Indeed, the CSE1-sg2 and CSE2-sg3 poplars had up to 25% higher saccharification efficiency than the WT control. Our results demonstrate that precise editing of CSE by CRISPR/Cas9 technology can improve lignocellulosic biomass without a growth penalty.

scholarly journals Patatin-Related Phospholipase AtpPLAIIIα Affects Lignification of Xylem in Arabidopsis and Hybrid Poplars

Plants ◽  
2020 ◽  
Vol 9 (4) ◽  
pp. 451 ◽  
Author(s):  
Jin Hoon Jang ◽  
Ok Ran Lee
Keyword(s):  
Cell Walls ◽  
Lignin Content ◽  
Lignin Biosynthesis ◽  
Biofuel Production ◽  
Amide Bonds ◽  
Hybrid Poplars ◽  
Plant Lipids ◽  
Lipid Acyl Hydrolase ◽  
Specific Regulation

Lipid acyl hydrolase are a diverse group of enzymes that hydrolyze the ester or amide bonds of fatty acid in plant lipids. Patatin-related phospholipase AIIIs (pPLAIIIs) are one of major lipid acyl hydrolases that are less closely related to potato tuber patatins and are plant-specific. Recently, overexpression of ginseng-derived PgpPLAIIIβ was reported to be involved in the reduced level of lignin content in Arabidopsis and the mature xylem layer of poplar. The presence of lignin-polysaccharides renders cell walls recalcitrant for pulping and biofuel production. The tissue-specific regulation of lignin biosynthesis, without altering all xylem in plants, can be utilized usefully by keeping mechanical strength and resistance to various environmental stimuli. To identify another pPLAIII homolog from Arabidopsis, constitutively overexpressed AtpPLAIIIα was characterized for xylem lignification in two well-studied model plants, Arabidopsis and poplar. The characterization of gene function in annual and perennial plants with respect to lignin biosynthesis revealed the functional redundancy of less lignification via downregulation of lignin biosynthesis-related genes.

scholarly journals Low Lignin Mutants and Reduction of Lignin Content in Grasses for Increased Utilisation of Lignocellulose

Agronomy ◽  
2019 ◽  
Vol 9 (5) ◽  
pp. 256 ◽  
Author(s):  
Cecilie S. L. Christensen ◽  
Søren K. Rasmussen
Keyword(s):  
Animal Feed ◽  
Lignin Content ◽  
Lignin Biosynthesis ◽  
Biofuel Production ◽  
Phenotypic Traits ◽  
Monolignol Biosynthesis ◽  
Genes Encoding ◽  
Regulated Gene Expression ◽  
Mutant Lines ◽  
The Impact

Biomass rich in lignocellulose from grasses is a major source for biofuel production and animal feed. However, the presence of lignin in cell walls limits its efficient utilisation such as in its bioconversion to biofuel. Reduction of the lignin content or alteration of its structure in crop plants have been pursued, either by regulating genes encoding enzymes in the lignin biosynthetic pathway using biotechnological techniques or by breeding naturally-occurring low lignin mutant lines. The aim of this review is to provide a summary of these studies, focusing on lignin (monolignol) biosynthesis and composition in grasses and, where possible, the impact on recalcitrance to bioconversion. An overview of transgenic crops of the grass family with regulated gene expression in lignin biosynthesis is presented, including the effect on lignin content and changes in the ratio of p-hydroxyphenyl (H), guaiacyl (G) and syringyl (S) units. Furthermore, a survey is provided of low-lignin mutants in grasses, including cereals in particular, summarising their origin and phenotypic traits together with genetics and the molecular function of the various genes identified.

scholarly journals Genetic, transcriptional, and regulatory landscape of monolignol biosynthesis pathway in Miscanthus × giganteus

2020 ◽  
Vol 13 (1) ◽  
Author(s):  
Xiaofei Zeng ◽  
Jiajing Sheng ◽  
Fenglin Zhu ◽  
Tianzi Wei ◽  
Lingling Zhao ◽  
...  
Keyword(s):  
Lignocellulosic Biomass ◽  
Lignin Content ◽  
Lignin Biosynthesis ◽  
Enzymatic Reactions ◽  
Biosynthesis Pathway ◽  
Miscanthus Giganteus ◽  
Monolignol Biosynthesis ◽  
Essential Knowledge ◽  
Main Component ◽  
Regulatory Landscape

Abstract Background Miscanthus × giganteus is widely recognized as a promising lignocellulosic biomass crop due to its advantages of high biomass production, low environmental impacts, and the potential to be cultivated on marginal land. However, the high costs of bioethanol production still limit the current commercialization of lignocellulosic bioethanol. The lignin in the cell wall and its by-products released in the pretreatment step is the main component inhibiting the enzymatic reactions in the saccharification and fermentation processes. Hence, genetic modification of the genes involved in lignin biosynthesis could be a feasible strategy to overcome this barrier by manipulating the lignin content and composition of M. × giganteus. For this purpose, the essential knowledge of these genes and understanding the underlying regulatory mechanisms in M. × giganteus is required. Results In this study, MgPAL1, MgPAL5, Mg4CL1, Mg4CL3, MgHCT1, MgHCT2, MgC3′H1, MgCCoAOMT1, MgCCoAOMT3, MgCCR1, MgCCR2, MgF5H, MgCOMT, and MgCAD were identified as the major monolignol biosynthetic genes in M. × giganteus based on genetic and transcriptional evidence. Among them, 12 genes were cloned and sequenced. By combining transcription factor binding site prediction and expression correlation analysis, MYB46, MYB61, MYB63, WRKY24, WRKY35, WRKY12, ERF021, ERF058, and ERF017 were inferred to regulate the expression of these genes directly. On the basis of these results, an integrated model was summarized to depict the monolignol biosynthesis pathway and the underlying regulatory mechanism in M. × giganteus. Conclusions This study provides a list of potential gene targets for genetic improvement of lignocellulosic biomass quality of M. × giganteus, and reveals the genetic, transcriptional, and regulatory landscape of the monolignol biosynthesis pathway in M. × giganteus.

scholarly journals Compositional analysis and characterization of lignocellulosic biomass from selected agricultural wastes

2020 ◽  
Vol 8 (1) ◽  
pp. 48-56
Author(s):  
Adewale Elijah Fadeyi ◽  
Saheed Olatunbosun Akiode ◽  
Stella A Emmanuel ◽  
Olajide Ebenezer Falayi
Keyword(s):  
Lignocellulosic Biomass ◽  
Lignin Content ◽  
Broad Band ◽  
Gravimetric Method ◽  
Compositional Analysis ◽  
Agricultural Wastes ◽  
Biofuel Production ◽  
Biomass Composition ◽  
Oh Groups

Agricultural wastes have been identified as a potential lignocellulosic biomass for bioethanol production. An accurate biomass characterization is needed to evaluate the new potential lignocelluloses biosource for biofuel production. This study evaluates the compositional analysis and characterization of three agricultural wastes (melon husk, moringa pod and mango endocarp). The samples were collected locally in Sheda Village, FCT, Abuja, Nigeria. The lignocellulose biomass composition of the samples was determined by using a proven economically viable gravimetric method and the samples were further characterized using the FTIR. The results showed that a significant amount of hemicelluloses content was found, from 19.38% to 27.74% and the highest amount was present in melon musk. The amount of cellulose ranging from 22.49% to 45.84% was found where the highest amount was found in mango endocarp. Lignin content was in the range of 22.62% to 29.87% and melon husk was shown to have the highest amount. The FTIR spectroscopic analysis showed a broad band at 3422.99 cm-1, 3422.66 cm-1, 3422.85 cm-1 (for mango endocarp, melon husk and moringa pod respectively) representing bonded –OH groups. The peak around 1637 cm-1 corresponds to C=C stretching of conjugated carboxylic acids. The aliphatic chains, -CH2- and –CH3, which form the basic structure of cellulose material, were seen at 1205.72, 1204.50 and 1206.24 cm-1. The signals at 1056.15, 1035.80 and 1055.86 cm-1 correspond to C-O-R (alcohols or esters) vibration. The results show that the samples contain significant quantity of lignocellulosic biomass. Thus, the agricultural wastes could be of valuable use in biofuel production.

scholarly journals RNAi suppression of lignin biosynthesis in sugarcane reduces recalcitrance for biofuel production from lignocellulosic biomass

2012 ◽  
Vol 10 (9) ◽  
pp. 1067-1076 ◽  
Author(s):  
Je Hyeong Jung ◽  
Walid M. Fouad ◽  
Wilfred Vermerris ◽  
Maria Gallo ◽  
Fredy Altpeter
Keyword(s):  
Lignocellulosic Biomass ◽  
Lignin Biosynthesis ◽  
Biofuel Production

Field-Grown Transgenic Hybrid Poplar with Modified Lignin Biosynthesis to Improve Enzymatic Saccharification Efficiency

2017 ◽  
Vol 5 (3) ◽  
pp. 2407-2414 ◽  
Author(s):  
Zhouyang Xiang ◽  
Suman Kumar Sen ◽  
Douyong Min ◽  
Dhanalekshmi Savithri ◽  
Fachuang Lu ◽  
...  
Keyword(s):  
Hybrid Poplar ◽  
Enzymatic Saccharification ◽  
Lignin Biosynthesis ◽  
Saccharification Efficiency ◽  
Modified Lignin

Forages for feedstocks of biorefineries in temperate environments: review of lignin research in bioenergy crops and some insight into Miscanthus studies

2014 ◽  
Vol 65 (11) ◽  
pp. 1199 ◽  
Author(s):  
Maria S. Dwiyanti ◽  
J. Ryan Stewart ◽  
Toshihiko Yamada
Keyword(s):  
Panicum Virgatum ◽  
Target Genes ◽  
Lignin Content ◽  
Genetic Regulation ◽  
Warm Season ◽  
Lignin Biosynthesis ◽  
Biofuel Production ◽  
Bioenergy Crops ◽  
Cost Efficient ◽  
Insight Into

Rhizomatous and perennial warm-season C4 grasses such as Miscanthus spp. and switchgrass (Panicum virgatum) are potential bioenergy crops for temperate regions. However, lignin in Miscanthus and switchgrass inhibits the cellulose digestion process during bioethanol production. One of the targets for improvement of forages from feedstocks to bioenergy crops is to develop a cost-efficient biorefinery process through lignin content manipulation. Numerous reports have shown that RNAi suppression of lignin-biosynthesis pathway genes can increase biomass fermentable sugar yields for biofuel production. These studies have also reported that RNAi suppression of cell-wall lignin biosynthesis can decrease biomass yield and resistance to biotic stress in the transgenic plants. Transcriptome and metabolome approaches can be used to clarify the networks and pathways of lignin biosynthesis to facilitate the identification of appropriate target genes for transformation. However, whole-genome sequencing of the forage species, which provides much-needed genomic information, is limited. Germplasm of natural, low-lignin mutants also plays a role in identification of genetic regulation of lignin content and this would be useful breeding material. Molecular markers have been developed and utilised to accelerate identification of quantitative trait loci/genes for traits relating to the biorefinery process. All of these studies will serve as basic information for supporting genetic improvement through classical breeding or genetic transformation, and offer the opportunity to develop cultivars which have enhanced biomass and are cost-efficient for biorefinery process.

scholarly journals The Osmotin-Like Protein Gene PdOLP1 Is Involved in Secondary Cell Wall Biosynthesis during Wood Formation in Poplar

2020 ◽  
Vol 21 (11) ◽  
pp. 3993 ◽  
Author(s):  
Shaofeng Li ◽  
Yaoxiang Zhang ◽  
Xuebing Xin ◽  
Changjun Ding ◽  
Fuling Lv ◽  
...  
Keyword(s):  
Cell Wall ◽  
Secondary Wall ◽  
Secondary Cell Wall ◽  
Vascular Tissue ◽  
Lignin Content ◽  
Hybrid Poplar ◽  
Lignin Biosynthesis ◽  
Negative Regulator ◽  
Cell Wall Biosynthesis ◽  
Secondary Cell Wall Biosynthesis

Osmotin-like proteins (OLPs) mediate defenses against abiotic and biotic stresses and fungal pathogens in plants. However, no OLPs have been functionally elucidated in poplar. Here, we report an osmotin-like protein designated PdOLP1 from Populus deltoides (Marsh.). Expression analysis showed that PdOLP1 transcripts were mainly present in immature xylem and immature phloem during vascular tissue development in P. deltoides. We conducted phenotypic, anatomical, and molecular analyses of PdOLP1-overexpressing lines and the PdOLP1-downregulated hybrid poplar 84K (Populus alba × Populus glandulosa) (Hybrid poplar 84K PagOLP1, PagOLP2, PagOLP3 and PagOLP4 are highly homologous to PdOLP1, and are downregulated in PdOLP1-downregulated hybrid poplar 84K). The overexpression of PdOLP1 led to a reduction in the radial width and cell layer number in the xylem and phloem zones, in expression of genes involved in lignin biosynthesis, and in the fibers and vessels of xylem cell walls in the overexpressing lines. Additionally, the xylem vessels and fibers of PdOLP1-downregulated poplar exhibited increased secondary cell wall thickness. Elevated expression of secondary wall biosynthetic genes was accompanied by increases in lignin content, dry weight biomass, and carbon storage in PdOLP1-downregulated lines. A PdOLP1 coexpression network was constructed and showed that PdOLP1 was coexpressed with a large number of genes involved in secondary cell wall biosynthesis and wood development in poplar. Moreover, based on transcriptional activation assays, PtobZIP5 and PtobHLH7 activated the PdOLP1 promoter, whereas PtoBLH8 and PtoWRKY40 repressed it. A yeast one-hybrid (Y1H) assay confirmed interaction of PtoBLH8, PtoMYB3, and PtoWRKY40 with the PdOLP1 promoter in vivo. Together, our results suggest that PdOLP1 is a negative regulator of secondary wall biosynthesis and may be valuable for manipulating secondary cell wall deposition to improve carbon fixation efficiency in tree species.

An eco-friendly biomass pretreatment strategy utilizing reusable enzyme mimicking nanoparticles for lignin depolymerization and biofuel production

2021 ◽  
Author(s):  
Rajiv CHANDRA RAJAK ◽  
Pathikrit Saha ◽  
Mamata S Singhvi ◽  
Darae Kwak ◽  
Danil Kim ◽  
...  
Keyword(s):  
Lignocellulosic Biomass ◽  
Biofuel Production ◽  
Biomass Pretreatment ◽  
Enzyme Mimicking ◽  
Lignin Depolymerization

Pretreatment of lignocellulosic biomass to specifically depolymerise lignin moieties without loss of carbohydrates as well as to minimize the generation of harmful intermediates during the process is a major challenge...

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