Author
Holly L. Baxter
Other affiliations: Oak Ridge National Laboratory
Bio: Holly L. Baxter is an academic researcher from University of Tennessee. The author has contributed to research in topics: Biomass & Lignin. The author has an hindex of 13, co-authored 24 publications receiving 778 citations. Previous affiliations of Holly L. Baxter include Oak Ridge National Laboratory.
Topics: Biomass, Lignin, Panicum virgatum, Bioenergy, Cellulose
Papers
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TL;DR: A versatile set of Gateway-compatible destination vectors (termed pANIC) was constructed to be used in monocot plants for transgenic crop improvement and was functionally validated in switchgrass and rice and allows for high-throughput screening of sequences of interest in othermonocot species as well.
Abstract: Summary Switchgrass (Panicum virgatum L.) is a C4 perennial grass and has been identified as a potential bioenergy crop for cellulosic ethanol because of its rapid growth rate, nutrient use efficiency and widespread distribution throughout North America. The improvement of bioenergy feedstocks is needed to make cellulosic ethanol economically feasible, and genetic engineering of switchgrass is a promising approach towards this goal. A crucial component of creating transgenic switchgrass is having the capability of transforming the explants with DNA sequences of interest using vector constructs. However, there are limited options with the monocot plant vectors currently available. With this in mind, a versatile set of Gateway- compatible destination vectors (termed pANIC) was constructed to be used in monocot plants for transgenic crop improvement. The pANIC vectors can be used for transgene overexpres- sion or RNAi-mediated gene suppression. The pANIC vector set includes vectors that can be utilized for particle bombardment or Agrobacterium-mediated transformation. All the vectors contain (i) a Gateway cassette for overexpression or silencing of the target sequence, (ii) a plant selection cassette and (iii) a visual reporter cassette. The pANIC vector set was function- ally validated in switchgrass and rice and allows for high-throughput screening of sequences of interest in other monocot species as well.
148 citations
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TL;DR: Downregulation of GAUT4 reduces HG and rhamnogalacturonan II (RGII), reduces wall calcium and boron, and increases extractability of cell wall sugars, which improves biomass yields and sugar release from biomass processing.
Abstract: Cell walls in crops and trees have been engineered for production of biofuels and commodity chemicals, but engineered varieties often fail multi-year field trials and are not commercialized. We engineered reduced expression of a pectin biosynthesis gene (Galacturonosyltransferase 4, GAUT4) in switchgrass and poplar, and find that this improves biomass yields and sugar release from biomass processing. Both traits were maintained in a 3-year field trial of GAUT4-knockdown switchgrass, with up to sevenfold increased saccharification and ethanol production and sixfold increased biomass yield compared with control plants. We show that GAUT4 is an α-1,4-galacturonosyltransferase that synthesizes homogalacturonan (HG). Downregulation of GAUT4 reduces HG and rhamnogalacturonan II (RGII), reduces wall calcium and boron, and increases extractability of cell wall sugars. Decreased recalcitrance in biomass processing and increased growth are likely due to reduced HG and RGII cross-linking in the cell wall.
118 citations
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TL;DR: The results of this study suggest that lignin down-regulation in switchgrass can confer real-world improvements in biofuel yield without negative consequences to biomass yield or disease susceptibility.
Abstract: Summary
Switchgrass (Panicum virgatum L.) is a leading candidate for a dedicated lignocellulosic biofuel feedstock owing to its high biomass production, wide adaptation and low agronomic input requirements. Lignin in cell walls of switchgrass, and other lignocellulosic feedstocks, severely limits the accessibility of cell wall carbohydrates to enzymatic breakdown into fermentable sugars and subsequently biofuels. Low-lignin transgenic switchgrass plants produced by the down-regulation of caffeic acid O-methyltransferase (COMT), a lignin biosynthetic enzyme, were analysed in the field for two growing seasons. COMT transcript abundance, lignin content and the syringyl/guaiacyl lignin monomer ratio were consistently lower in the COMT-down-regulated plants throughout the duration of the field trial. In general, analyses with fully established plants harvested during the second growing season produced results that were similar to those observed in previous greenhouse studies with these plants. Sugar release was improved by up to 34% and ethanol yield by up to 28% in the transgenic lines relative to controls. Additionally, these results were obtained using senesced plant material harvested at the end of the growing season, compared with the young, green tissue that was used in the greenhouse experiments. Another important finding was that transgenic plants were not more susceptible to rust (Puccinia emaculata). The results of this study suggest that lignin down-regulation in switchgrass can confer real-world improvements in biofuel yield without negative consequences to biomass yield or disease susceptibility.
105 citations
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TL;DR: This review will recapitulate the knowledge of lignin biosynthesis and discuss how lign in has been modified and the consequences for the host plant.
Abstract: Lignocellulosic feedstocks can be converted to biofuels, which can conceivably replace a large fraction of fossil fuels currently used for transformation. However, lignin, a prominent constituent of secondary cell walls, is an impediment to the conversion of cell walls to fuel: the recalcitrance problem. Biomass pretreatment for removing lignin is the most expensive step in the production of lignocellulosic biofuels. Even though we have learned a great deal about the biosynthesis of lignin, we do not fully understand its role in plant biology, which is needed for the rational design of engineered cell walls for lignocellulosic feedstocks. This review will recapitulate our knowledge of lignin biosynthesis and discuss how lignin has been modified and the consequences for the host plant.
97 citations
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TL;DR: Transient expression was increased by double or triple wounding treatments such as mixing by vortex with carborundum, sonication, separation by centrifugation, and heat shock, and the addition of thiol compounds during cocultivation increased transient expression levels from 6% to 54%.
Abstract: Background
Grasses are relatively recalcitrant to genetic transformation in comparison to certain dicotyledons, yet they constitute some of the most important biofuel crops. Genetic transformation of switchgrass (Panicum virgatum L.) has previously been reported after cocultivation of explants with Agrobacterium and biolistics of embryogenic calli. Experiments to increase transient gene expression in planta may lead to stable transformation methods with increased efficiency.
75 citations
Cited by
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01 Jul 2004
TL;DR: In this article, the authors developed a center to address state-of-the-art research, create innovating educational programs, and support technology transfers using commercially viable results to assist the Army Research Laboratory to develop the next generation Future Combat System in the telecommunications sector that assures prevention of perceived threats, and non-line of sight/Beyond line of sight lethal support.
Abstract: Home PURPOSE OF THE CENTER: To develop the center to address state-of-the-art research, create innovating educational programs, and support technology transfers using commercially viable results to assist the Army Research Laboratory to develop the next generation Future Combat System in the telecommunications sector that assures prevention of perceived threats, and Non Line of Sight/Beyond Line of Sight lethal support.
1,713 citations
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TL;DR: This review provides a “beginning‐to‐end” analysis of the recent advances reported in lignin valorisation, with particular emphasis on the improved understanding of lign in's biosynthesis and structure.
Abstract: 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.
1,390 citations
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TL;DR: It is demonstrated that the class 2 type VI RNA-guided RNA-targeting CRISPR–Cas effector Cas13a (previously known as C2c2) can be engineered for mammalian cell RNA knockdown and binding and is established as a flexible platform for studying RNA in mammalian cells and therapeutic development.
Abstract: RNA has important and diverse roles in biology, but molecular tools to manipulate and measure it are limited. For example, RNA interference can efficiently knockdown RNAs, but it is prone to off-target effects, and visualizing RNAs typically relies on the introduction of exogenous tags. Here we demonstrate that the class 2 type VI RNA-guided RNA-targeting CRISPR-Cas effector Cas13a (previously known as C2c2) can be engineered for mammalian cell RNA knockdown and binding. After initial screening of 15 orthologues, we identified Cas13a from Leptotrichia wadei (LwaCas13a) as the most effective in an interference assay in Escherichia coli. LwaCas13a can be heterologously expressed in mammalian and plant cells for targeted knockdown of either reporter or endogenous transcripts with comparable levels of knockdown as RNA interference and improved specificity. Catalytically inactive LwaCas13a maintains targeted RNA binding activity, which we leveraged for programmable tracking of transcripts in live cells. Our results establish CRISPR-Cas13a as a flexible platform for studying RNA in mammalian cells and therapeutic development.
1,274 citations
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TL;DR: The goal of this study is to review the fundamental structures and chemistries of wood and wood-derived materials, which are essential for a wide range of existing and new enabling technologies.
Abstract: 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...
1,031 citations
05 Mar 2001
TL;DR: It is indicated that lignin and cellulose deposition could be regulated in a compensatory fashion, which may contribute to metabolic flexibility and a growth advantage to sustain the long-term structural integrity of woody perennials.
Abstract: Because lignin limits the use of wood for fiber, chemical, and energy production, strategies for its downregulation are of considerable interest. We have produced transgenic aspen (Populus tremuloides Michx.) trees in which expression of a lignin biosynthetic pathway gene Pt4CL1 encoding 4-coumarate:coenzyme A ligase (4CL) has been downregulated by antisense inhibition. Trees with suppressed Pt4CL1 expression exhibited up to a 45% reduction of lignin, but this was compensated for by a 15% increase in cellulose. As a result, the total lignin–cellulose mass remained essentially unchanged. Leaf, root, and stem growth were substantially enhanced, and structural integrity was maintained both at the cellular and whole-plant levels in the transgenic lines. Our results indicate that lignin and cellulose deposition could be regulated in a compensatory fashion, which may contribute to metabolic flexibility and a growth advantage to sustain the long-term structural integrity of woody perennials.
717 citations