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At least 37 records · Page 2

W Photoprotection in Tropical Marine Organisms

Increasing levels of ultraviolet (UV) radiation reaching the earth's surface which results from stratospheric ozone depletions could have serious implications for terrestrial plants and for aquatic organisms within the euphotic zone. A documented 9% decline in ozone at mid-latitudes is considered to produce a 12% increase in harmful UV radiation. The biologically damaging effects of higher UV levels, particularly W-B (280-320 rim), could manifest earlier in the tropics because of the relative thinness of the earth's equatorial ozone layer. Tropical marine organisms are also living close to their upper tolerance levels of water temperature, However, despite the large potential effects on plants and animals, little is known about UV effects on tropical ecosystems. Long-term ecological studies are needed to quantify the effects of increased UV radiation on terrestrial and marine ecosystems and to produce reliable data for prediction. Plants have developed several mechanisms to protect themselves from harmful UV radiation, one of which is the production of secondary leaf pigments that absorb W-B radiation (screening pigments). A higher concentration of screening pigments (e.g. flavonoids) in leaves may be interpreted as a natural response to increased W radiation. If higher concentrations of flavonoids filter out the excessive W radiation, no damage will occur, as suggested by Caldwell et al. (1989) and Tevini (1993). Failure to screen all W-B may result in deleterious effects on photosynthesis, plant genetic material, and plant and leaf morphology and growth. Eventually this will have an impact on ecosystem processes, structure, species composition, and productivity. This paper describes an ongoing project that is assessing the responses of mangroves, seagrasses and corals to W radiation by studying pigment concentrations, biophysical parameters, and variations in spectral reflectance in the field and in W-reduction experiments. Preliminary results on the distribution of W-absorbing flavonoid compounds in red mangroves (Rhizophora mangle) and the seagrass Thalassia testudinum, are presented. This research also provides, for the first time, a permanent record of daily W irradiance measurements at a tropical -location.

Armstrong, Roy A.↗

Engineered Sorghum Bagasse Enables a Sustainable Biorefinery with p-Hydroxybenzoic Acid-Based Deep Eutectic Solvent

Integrating multidisciplinary research in plant genetic engineering and renewable deep eutectic solvents (DESs) can facilitate a sustainable and economic biorefinery. In this work, we leveraged a plant genetic engineering approach to specifically incorporate C6C1 monomers into the lignin structure. By expressing the bacterial ubiC gene in sorghum, p-hydroxybenzoic acid (PB)-rich lignin was incorporated into the plant cell wall while this monomer was completely absent in the lignin of the wild-type (WT) biomass. A DES was synthesized with choline chloride (ChCl) and PB and applied to the pretreatment of the PB-rich mutant biomass for a sustainable biorefinery. The release of fermentable sugars was significantly enhanced (~190 % increase) compared to untreated biomass by the DES pretreatment. In particular, the glucose released from the pretreated mutant biomass was up to 12 % higher than that from the pretreated WT biomass. Lignin was effectively removed from the biomass with the preservation of more than half of the β-Ο-4 linkages without condensed aromatic structures. Hydrogenolysis of the fractionated lignin was conducted to demonstrate the potential of phenolic compound production. In addition, a simple hydrothermal treatment could selectively extract PB from the same engineered lignin, showing a possible circular biorefinery. These results suggest that the combination of PB-based DES and engineered PB-rich biomass is a promising strategy to achieve a sustainable closed-loop biorefinery.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗

Methods for controlling cell wall biosynthesis and genetically modified plants

The present disclosure provides methods of producing plants with preferred levels of cell wall biosynthesis; and uses of such plants. The inventors have identified that the GFR9, CCoAOMT and MYB41 genes are major regulators of the cell wall biosynthesis pathway. Plants with modulated cell wall biosynthesis, based on modulation of the expression or activity of the GFR9, CCoAOMT and MYB41 genes, have divergent uses including pulp and paper production, and bioproduct production.

Tschaplinski, Timothy J.↗

Engineering plants for spaceflight environments

The conversion efficiency of radiation into biomass and yield has steadily increased for centuries because of continued improvement in both plant genetics and environmental control. Considerable effort has gone into improving the environment for plant growth in space, but work has only begun to engineer plants for spaceflight. Genetic manipulation offers tremendous potential to improve our ability to study gravitational effects. Genetic manipulation will also be necessary to build an efficient regenerative life support system. We cannot fully characterize plant response to the spaceflight environment without understanding and manipulating their genetic composition. Identification and selection of the existing germplasm is the first step. There are thousands of cultivars of each of our major crop plants, each specifically adapted to a unique environment on our planet. Thousands of additional lines are held in national germplasm collections to maintain genetic diversity. Spaceflight imposes the need to tap this diversity. Existing lines need to be evaluated in the environment that is characteristic of closed-system spaceflight conditions. Many of the plant growth challenges we confront in space can be better solved through genetic change than by hardware engineering. Ten thousand years of plant breeding has demonstrated the value of matching genetics with the environment. For example, providing continuous light can increase plant growth in space, but this often induces calcium deficiencies because Ca is not supplied by guttation during a dark period. This deficiency cannot be eliminated through increased root-zone and foliar Ca applications. It can be solved, in wheat, through genetic selection of lines that do not have the deficiency. Subsequent comparison of lines with and without the Ca deficiency has also helped us understand the nature of the problem.

Non-NASA Center↗

Validation of a metabolite–GWAS network for Populus trichocarpa family 1 UDP-glycosyltransferases

Metabolite genome-wide association studies (mGWASs) are increasingly used to discover the genetic basis of target phenotypes in plants such as Populus trichocarpa , a biofuel feedstock and model woody plant species. Despite their growing importance in plant genetics and metabolomics, few mGWASs are experimentally validated. Here, we present a functional genomics workflow for validating mGWAS-predicted enzyme–substrate relationships. We focus on uridine diphosphate–glycosyltransferases (UGTs), a large family of enzymes that catalyze sugar transfer to a variety of plant secondary metabolites involved in defense, signaling, and lignification. Glycosylation influences physiological roles, localization within cells and tissues, and metabolic fates of these metabolites. UGTs have substantially expanded in P. trichocarpa , presenting a challenge for large-scale characterization. Using a high-throughput assay, we produced substrate acceptance profiles for 40 previously uncharacterized candidate enzymes. Assays confirmed 10 of 13 leaf mGWAS associations, and a focused metabolite screen demonstrated varying levels of substrate specificity among UGTs. A substrate binding model case study of UGT-23 rationalized observed enzyme activities and mGWAS associations, including glycosylation of trichocarpinene to produce trichocarpin, a major higher-order salicylate in P. trichocarpa. We identified UGTs putatively involved in lignan, flavonoid, salicylate, and phytohormone metabolism, with potential implications for cell wall biosynthesis, nitrogen uptake, and biotic and abiotic stress response that determine sustainable biomass crop production. Our results provide new support for in silico analyses and evidence-based guidance for in vivo functional characterization.

59 BASIC BIOLOGICAL SCIENCES↗

Understanding Plant Signaling Via Innovations in Probe Delivery and Imaging

The goal of this project was to develop tools for studying plant processes in living tissues. We wanted a method for introducing probes for studying plant processes such that plants could remain intact and imaged iteratively. Towards this end, we successfully optimized carbon nanofibers that we showed could deliver non-permeable signaling probes/biomolecules to plant cells. These were created on a rigid backing and also successfully transferred to a flexible backing to use with curved plant structures. We also built fiber optic microscopes that permit iterative non-destructive fluorescence or light micrographs and created protocols and devices to stabilize plant organs during imaging. These tools were developed in conjunction with research aimed understanding receptor-mediated peptide trafficking and responses relevant to plant cell growth and defense tradeoff. During this project, we designed and built two fiberoptic fluorescence microscopes; (b) designed, fabricated and validated different nanospike designs for delivering probes to plants, including the feasibility of delivering DNA constructs for expression studies; (c) developed fluorescent peptide probes (active and inactive versions of a secreted receptor ligand called phytosulfokine or PSK) and performed mobility tests after application to plants using microscopy; (d) used appropriate genetic plant backgrounds to facilitate PSK trafficking and response mechanisms; (e) determined the transcriptional changes due to PSK-induced signaling related to growth/defense tradeoff; (f) performed experiments showing the growth/defense tradeoff involves repression of specific defense components in response to PSK. This project also involved the training of two PhD students and five Masters students.

47 OTHER INSTRUMENTATION↗

Research for Sustainable Bioenergy: Linking Genomic and Ecosystem Sciences, Workshop, October 2-4, 2013 Report

The design of sustainable biofuel systems requires knowledge about key plant-microbe-environment interactions that provide a range of ecosystem services. Most critical is a mechanistic understanding of how candidate biofuel plants interact with biotic and abiotic factors to affect the ecosystem outcomes that define sustainability. Recent advances in the genomic sciences can contribute immensely to the knowledge needed to design such systems. For example, progress in plant genomics will enable the inclusion of sustainability traits in future feedstocks, and advances in microbial genomics will allow insights into plantmicrobe-soil interactions that might be used to manage and support plant productivity and vigor. Linking these advances to breakthroughs in ecosystem science enables the use of systems biology in the fundamental design of sustainable biofuel systems. To identify research opportunities in developing such systems, the Department of Energy’s (DOE) Office of Biological and Environmental Research (BER) held the Research for Sustainable Bioenergy Workshop on Oct. 2–4, 2013, in Germantown, Maryland. The workshop convened more than 30 researchers with a broad and diverse range of expertise, including ecology, microbiology, plant genetics, genomics, computational biology, and modeling. Participants discussed and identified research gaps, challenges, and opportunities for enhancing the understanding of influences that biotic, abiotic, and genetic factors have on long-term plant feedstock performance and the delivery of ecosystem services at multiple scales. This report identifies the key topics and questions that could be addressed effectively to achieve this understanding. Research opportunities are organized into four categories: (1) plant systems, encompassing plant productivity, resource use efficiency, genotype/phenotype breeding, and crop diversity; (2) the plant microbiome, which includes microbes living in close association within or adjacent to plants; (3) ecosystem processes, such as carbon capture, greenhouse gas mitigation, and hydrologic processes; and (4) multiscale modeling, which integrates and extends results across spatiotemporal scales.

59 BASIC BIOLOGICAL SCIENCES↗

A Novel Gene Stacking Method in Plant Transformation Utilizing Split Selectable Markers

Gene stacking, the process of introducing multiple genes into a single plant to enhance desired traits, is essential for plant genetic improvement through both conventional breeding and genetic transformation. In general, transformation-based gene stacking can be achieved through either co-transformation to simultaneously introduce multiple genes or sequential multi-round transformation. While co-transformation is generally faster and more efficient than sequential multi-round transformation, it often requires two selectable marker genes, which confer resistance to antibiotics, for selecting transgenic events. However, in most cases, there is only one best selectable marker gene for a specific plant species or genotype. Also, it is harder to optimize the concentrations of two antibiotics for co-transformation than using one antibiotic for selecting transgenic events. To overcome this challenge, we recently developed an innovative split selectable marker system for plant co-transformation, allowing the use of one selectable marker gene to select transgenic events. This method involves constructing two binary vectors, each carrying a subset of genes of interest and a partial fragment of the selectable marker gene, which is connected to a partial intein fragment. Following Agrobacterium -mediated co-transformation, plants harboring both binary vectors are selected using a single antibiotic, such as kanamycin. This split-marker system can be used to co-transform multiple genes into both herbaceous and woody plants, accelerating genetic improvement of polygenic traits or integrative improvement of multiple traits to simultaneously increase crop yield and quality.

59 BASIC BIOLOGICAL SCIENCES↗

A high-throughput skim-sequencing approach for genotyping, dosage estimation and identifying translocations

The development of next-generation sequencing (NGS) enabled a shift from array-based genotyping to directly sequencing genomic libraries for high-throughput genotyping. Even though whole-genome sequencing was initially too costly for routine analysis in large populations such as breeding or genetic studies, continued advancements in genome sequencing and bioinformatics have provided the opportunity to capitalize on whole-genome information. As new sequencing platforms can routinely provide high-quality sequencing data for sufficient genome coverage to genotype various breeding populations, a limitation comes in the time and cost of library construction when multiplexing a large number of samples. Here we describe a high-throughput whole-genome skim-sequencing (skim-seq) approach that can be utilized for a broad range of genotyping and genomic characterization. Using optimized low-volume Illumina Nextera chemistry, we developed a skim-seq method and combined up to 960 samples in one multiplex library using dual index barcoding. With the dual-index barcoding, the number of samples for multiplexing can be adjusted depending on the amount of data required, and could be extended to 3,072 samples or more. Panels of doubled haploid wheat lines ( Triticum aestivum , CDC Stanley x CDC Landmark), wheat-barley ( T . aestivum x Hordeum vulgare ) and wheat-wheatgrass ( Triticum durum x Thinopyrum intermedium ) introgression lines as well as known monosomic wheat stocks were genotyped using the skim-seq approach. Bioinformatics pipelines were developed for various applications where sequencing coverage ranged from 1 × down to 0.01 × per sample. Using reference genomes, we detected chromosome dosage, identified aneuploidy, and karyotyped introgression lines from the skim-seq data. Leveraging the recent advancements in genome sequencing, skim-seq provides an effective and low-cost tool for routine genotyping and genetic analysis, which can track and identify introgressions and genomic regions of interest in genetics research and applied breeding programs.

60 APPLIED LIFE SCIENCES↗

Large-scale replicated field study of maize rhizosphere identifies heritable microbes

Up until now, soil microbes that colonize plant roots and are responsive to differences in plant genotype remain to be ascertained for agronomically important crops. From a very large-scale longitudinal field study of 27 maize inbred lines planted in three fields, with partial replication 5 y later, we identify root-associated microbiota exhibiting reproducible associations with plant genotype. Analysis of 4,866 samples identified 143 operational taxonomic units (OTUs) whose variation in relative abundances across the samples was significantly regulated by plant genotype, and included five of seven core OTUs present in all samples. Plant genetic effects were significant amid the large effects of plant age on the rhizosphere microbiome, regardless of the specific community of each field, and despite microbiome responses to climate events. Seasonal patterns showed that the plant root microbiome is locally seeded, changes with plant growth, and responds to weather events. However, against this background of variation, specific taxa responded to differences in host genotype. If shown to have beneficial functions, microbes may be considered candidate traits for selective breeding.

59 BASIC BIOLOGICAL SCIENCES↗

Agrobacterium tumefaciens : A Bacterium Primed for Synthetic Biology

Agrobacterium tumefaciens is an important tool in plant biotechnology due to its natural ability to transfer DNA into the genomes of host plants. Genetic manipulations of A. tumefaciens have yielded considerable advances in increasing transformational efficiency in a number of plant species and cultivars. Moreover, there is overwhelming evidence that modulating the expression of various mediators of A. tumefaciens virulence can lead to more successful plant transformation; thus, the application of synthetic biology to enable targeted engineering of the bacterium may enable new opportunities for advancing plant biotechnology. In this review, we highlight engineering targets in both A. tumefaciens and plant hosts that could be exploited more effectively through precision genetic control to generate high-quality transformation events in a wider range of host plants. We then further discuss the current state of A. tumefaciens and plant engineering with regard to plant transformation and describe how future work may incorporate a rigorous synthetic biology approach to tailor strains of A. tumefaciens used in plant transformation.

59 BASIC BIOLOGICAL SCIENCES↗

Local and latitudinal variation in abundance: the mechanisms shaping the distribution of an ecosystem engineer

Ecological processes that determine the abundance of species within ecological communities vary across space and time. These scale-dependent processes are especially important when they affect key members of a community, such as ecosystem engineers that create shelter and food resources for other species. Yet, few studies have examined the suite of processes that shape the abundance of ecosystem engineers. Here, we evaluated the relative influence of temporal variation, local processes, and latitude on the abundance of an engineering insect—a rosette-galling midge, Rhopalomyia solidaginis (Diptera: Cecidomyiidae). Over a period of 3–5 years, we studied the density and size of galls across a suite of local experiments that manipulated genetic variation, soil nutrient availability, and the removal of other insects from the host plant, Solidago altissima (tall goldenrod). We also surveyed gall density within a single growing season across a 2,300 km latitudinal transect of goldenrod populations in the eastern United States. At the local scale, we found that host-plant genotypic variation was the best predictor of rosette gall density and size within a single year. We found that the removal of other insect herbivores resulted in an increase in gall density and size. The amendment of soil nutrients for four years had no effect on gall density, but galls were smaller in carbon-added plots compared to control and nitrogen additions. Finally, we observed that gall density varied several fold across years. At the biogeographic scale, we observed that the density of rosette gallers peaked at mid-latitudes. Using meta-analytic approaches, we found that the effect size of time, followed by host-plant genetic variation and latitude were the best predictors of gall density. Taken together, our study provides a unique comparison of multiple factors across different spatial and temporal scales that govern engineering insect herbivore density.

54 ENVIRONMENTAL SCIENCES↗

Association mapping across a multitude of traits collected in diverse environments in maize

Abstract Classical genetic studies have identified many cases of pleiotropy where mutations in individual genes alter many different phenotypes. Quantitative genetic studies of natural genetic variants frequently examine one or a few traits, limiting their potential to identify pleiotropic effects of natural genetic variants. Widely adopted community association panels have been employed by plant genetics communities to study the genetic basis of naturally occurring phenotypic variation in a wide range of traits. High-density genetic marker data—18M markers—from 2 partially overlapping maize association panels comprising 1,014 unique genotypes grown in field trials across at least 7 US states and scored for 162 distinct trait data sets enabled the identification of of 2,154 suggestive marker-trait associations and 697 confident associations in the maize genome using a resampling-based genome-wide association strategy. The precision of individual marker-trait associations was estimated to be 3 genes based on a reference set of genes with known phenotypes. Examples were observed of both genetic loci associated with variation in diverse traits (e.g., above-ground and below-ground traits), as well as individual loci associated with the same or similar traits across diverse environments. Many significant signals are located near genes whose functions were previously entirely unknown or estimated purely via functional data on homologs. This study demonstrates the potential of mining community association panel data using new higher-density genetic marker sets combined with resampling-based genome-wide association tests to develop testable hypotheses about gene functions, identify potential pleiotropic effects of natural genetic variants, and study genotype-by-environment interaction.

59 BASIC BIOLOGICAL SCIENCES↗

A century of genetic variation inferred from a persistent soil‐stored seed bank

Abstract Stratigraphic accretion of dormant propagules in soil can result in natural archives useful for studying ecological and evolutionary responses to environmental change. Few attempts have been made, however, to use soil‐stored seed banks as natural archives, in part because of concerns over nonrandom attrition and mixed stratification. Here, we examine the persistent seed bank of Schoenoplectus americanus , a foundational brackish marsh sedge, to determine whether it can serve as a resource for reconstructing historical records of demographic and population genetic variation. After assembling profiles of the seed bank from radionuclide‐dated soil cores, we germinated seeds to “resurrect” cohorts spanning the 20th century. Using microsatellite markers, we assessed genetic diversity and differentiation among depth cohorts, drawing comparisons to extant plants at the study site and in nearby and more distant marshes. We found that seed density peaked at intermediate soil depths. We also detected genotypic differences among cohorts as well as between cohorts and extant plants. Genetic diversity did not decline with depth, indicating that the observed pattern of differentiation is not due to attrition. Patterns of differentiation within and among extant marshes also suggest that local populations persist as aggregates of small clones, likely reflecting repeated seedling recruitment and low immigration from admixed regional gene pools. These findings indicate that persistent and stratified soil‐stored seed banks merit further consideration as resources for reconstructing decadal‐ to century‐long records that can lend insight into the tempo and nature of ecological and evolutionary processes that shape populations over time.

54 ENVIRONMENTAL SCIENCES↗

Feed Your Friends: Do Plant Exudates Shape the Root Microbiome?

Plant health in natural environments depends on interactions with complex and dynamic communities comprising macro- and microorganisms. While many studies have provided insights into the composition of rhizosphere microbiomes (rhizobiomes), little is known about whether plants shape their rhizobiomes. In this work, we discuss physiological factors of plants that may govern plant–microbe interactions, focusing on root physiology and the role of root exudates. Given that only a few plant transport proteins are known to be involved in root metabolite export, we suggest novel families putatively involved in this process. Finally, building off of the features discussed in this review, and in analogy to well-known symbioses, we elaborate on a possible sequence of events governing rhizobiome assembly. Recent advances in sequencing technology have been enabling high-throughput characterization of highly complex plant-associated microbial communities, which are relevant for plant health. Metagenomic approaches have been identifying the metabolic potential of microbes, and are starting to reveal functional groups of microbes, reducing the overall complexity of rhizobiomes. Metabolomic studies are uncovering differential root exudation in various environments and plant developmental stages, as well as consumption of specific exometabolites by microbes. Genome-wide association studies and other comparative genomic approaches have been revealing a link between plant genetic factors, exudation profiles, and microbiome compositions. Finally, molecular studies have been characterizing plant transport proteins necessary for interactions with specific microbes, although the transport of most nutrients and signaling molecules remains unexplored.

59 BASIC BIOLOGICAL SCIENCES↗

Comprehensive SNP Data for 1,323 GWAS Population in Populus trichocarpa and Combined Annotation Files for P. trichocarpa v3.0 and v3.1

The VCF dataset includes genetic variations found in 1,323 Populus trichocarpa genotypes, providing valuable information for scientists studying plant genetics. Researchers have generated this dataset using whole-genome DNA short-read sequencing on the Illumina Genome Analyzer, HiSeq 2000, and HiSeq 2500 platforms. This sequencing effort ensured a minimum expected sequencing depth of 15×. The dataset comprises more than 9.7 million single nucleotide polymorphisms (SNPs) and indel variants. The combined annotation files are derived from P. trichocarpa v3.0 and v3.1. We merged these files to create a comprehensive annotation file used for GWAS analysis. In total, 38,830 genes overlapped between the two versions. For overlapping genes, we defined the start as the smaller and the end as the larger among the two versions to increase the likelihood of locating candidate genetic loci. Additionally, we included 2,505 unique genes from v3.0 and 4,120 unique genes from v3.1, resulting in a total of 45,455 genes in the updated annotation file.

09 BIOMASS FUELS↗