Lawrence B. Smart1, Craig H. Carlson1, Dustin Wilkerson1, Chase Crowell2, Brennan Hyden1, Fred E. Gouker1, Christine D. Smart2, Yongwook Choi3, Agnes P. Chan3, Christopher D. Town3, Ran Zhou4, Stephen P. DiFazio4, Matt Olson5, Ken Keefover-Ring6, Jeremy Schmutz7
1 Horticulture Section, Plant Pathology and Plant-Microbe Biology Section2, School of Integrative Plant Sciences, Cornell University, Geneva, NY USA
3 Plant Genomics Group, J. Craig Venter Institute, Rockville, MD USA
4Dept. of Biology, West Virginia University, Morgantown, WV USA
5Dept. of Biological Sciences, Texas Tech University, Lubbock, TX USA
6Botany Dept., University of Wisconsin, Madison, WI USA
7Hudson Alpha Institute, Huntsville, AL USA
Shrub willow is among the best suited perennial crops for bioenergy feedstock production in the Northeast US due to its high yields, ease of conversion, and overall sustainability. Breeding efforts are aimed at improving yields on typical ag land, strengthening pest and disease resistance, and developing cultivars that can produce viable yields on highly disturbed sites, such as reclaimed mine land. In order to map resistance to willow leaf rust and other traits of interest including yield components, insect resistance, and physiological traits, we have developed three mapping resources: a small association panel planted on three sites, an F2 mapping population of Salix purpurea with 485 progeny planted in Geneva, NY, and eight species hybrid F1 mapping populations with a common parent of S. purpurea. Each of these F1 hybrid families has either S. purpurea 94006 as the female parent (the reference genome) or S. purpurea 94001 as the male parent crossed to individuals of S. suchowensis, S. viminalis, S. udensis, S. integra, S. koriyanagi, or S. alberti and is planted adjacent to the F2 population. In collaboration with the Dept of Energy Joint Genome Institute, we have developed high quality reference genomes of both male and female S. purpurea and have characterized the sex determination regions of the Z and W versions of Chr15. While there are long-standing models to describe the genetic basis for heterosis (hybrid vigor) in inbred crops, none of these can be directly applied to explain heterosis in undomesticated, outcrossing, highly heterozygous, and polyploid species, such as in Salix bioenergy crops. We produced a series of reciprocal crosses to test for heterosis in intra-specific F1 and F2 families of Salix purpurea, inter-specific diploid F1 hybrids between S. purpurea and S. viminalis, and inter-specific F1 hybrids between the diploids S. purpurea or S. viminalis and the tetraploid S. miyabeana. Non-additive inheritance patterns (principally dominant inheritance) account for a majority of the differential expression observed in all families assayed. Sex-biased gene expression also contributes to non-additive inheritance of expression and reveals the genetic basis for dimorphism associated with dioecy.
Stéphane Maury*1, Régis Fichot1, Mamadou Dia Sow1, Alain Delaunay1, Isabelle Le Jan1, Gwenvaël Laskar1, Marie-Claude Lesage-Descause2, Corinne Buret2, Vanina Guerin2, Odile Rogier2, Marie-Anne Lelu-Walter2, Vincent Ségura2, Grégoire Le Provost3, François Ehrenmann3, Ludovic Duvaux3, Isabelle Lesur3, Erwan Guichoux3, Christophe Plomion3, Peter Civan4, Jérome Salse4, Christophe Ambroise5, Sveltlana Gribkova5, Jorg Tost6, Jean-François Trontin7, Isabel Allona8, Steven Strauss9
1 Laboratoire de Biologie des Ligneux et des Grandes Cultures, LBLGC, 1207 USC1328, INRA-Université Orléans, Orléans FRANCE
2 Biologie intégrée pour la valorisation de la diversité des arbres et de la forêt, BioForA, UMR 0588, INRA-ONF, Orléans FRANCE
3 Biodiversité, Gènes et Communautés, BIOGECO, UMR 1202, INRA-Université Bordeaux, Bordeaux FRANCE
4 Génétique, Diversité et Écophysiologie des Céréales, GDEC, UMR 1095, INRA-Université Clermont Auvergne, Clermont-Ferrand FRANCE
5 Laboratoire de Mathématiques et Modélisation d'Évry, LaMME, UMR 8071, CNRS-Université d'Evry Val d'Essonne, Evry FRANCE
6 Laboratoire Épigénétique et Environnement, LEE, CNRGH CEA, Evry FRANCE)
7 FCBA, Pôle Biotechnologies et Sylviculture avancée, Cestas, FRANCE
8 Centro de Biotecnología y Genómica de Plantas, Universidad Politécnica de Madrid (UPM) 28223 Madrid, SPAIN
9 Department of Forest Ecosystems and Society, Oregon State University, Corvallis, OR 97331, USA
*Author for correspondence: stephane.maury@univ-orleans.fr
Ongoing global climate changes in progress will impact forest productivity notably through reduced water availability and heat periods. One possibility to adapt is phenotypic plasticity for which epigenetic mechanisms are proposed to be a main source of flexibility. Our objective is to evaluate the potential of epigenetics and more specifically DNA methylation to significantly participate to phenotypic plasticity in trees in response to drought or heat. Over the 10 last years, using an integrative approach with ecophysiological, biochemical, transcriptomics, epigenomics (MeDIP, WGBS, Mobilome) and reverse genetics (RNAi lines) tools, we were able to dissect in the shoot apical meristem (center of the shoot morphogenesis) or somatic embryos the response of trees to environmental variations notably in Salicacae such as poplar. This work was assessed in distinct experimental set-ups from in vitro, greenhouse to field plantations as well as during the stress or months post-stress. Our recent data (Lafon-Placette et al., 2018; Le Gac et al., 2018; Sow et al., 2018a; Sow et al., 2018b, Le Gac et al., 2019; Maury et al., 2019 and unpublished data) showed that Differentially Methylated Regions (DMRs) are associated to active TEs and differentially expressed genes with biological functions related to phytohormone signaling and may participate to an environmental epigenetic memory. Altogether, our data proposed that DNA methylation is a source of flexibility associated to phenotypic plasticity in trees opening perspectives for tree management. The role of epigenetic mechanisms in tree adaptation and microevolution will be also presented in the frame of the project EPITREE 2018-2021 (ANR-17-CE32-0009-01, https://www6.inra.fr/epitree-project_eng/The-Epitree-project).
Ben Bubner1, Volker Schneck1, Matthias Zander2, Jan Gloger2, Christian Ulrichs2
1Thünen Institute of Forest Genetics, Federal Research Institute for Rural Areas, Forestry and Fisheries, Eberswalder Chaussee 3A 15377 Waldsieversdorf
2 Humboldt-Universität zu Berlin, Faculty of Life Sciences, Albrecht Daniel Thaer-Institute of Agricultural and Horticultural Sciences, Division Urban Plant Ecophysiology55/ 57 14195-Berlin
Willows can be planted in an environmental context (e.g. protection of river embankments, early spring nutrient delivery for bees) and in an economic context. In commercial plantations biomass willows are used for fuel production (heating) in in short rotation coppices (SRC). The clones available for SRC plantations in Europe are based on Salix viminalis and relative species. Other potential commercial uses of willows are the extraction of natural salicylic compounds as alternatives to chemically synthesized acetyl salicylic acid (medical use) and the extraction of natural phenolic compounds from the cortex as alternative to mineral tanning agents (leather production).
In order to provide a wider genetic base, a collection of several hundred wild type genotypes of S. daphnoides, S. purpurea and S. pentandra was used for an extensive crossing and selection program. Wild type clones and crossings have been selected for several parameters: drought stress resistance as tested in greenhouse experiments with controlled irrigation, salicylate content measured by HPLC, biomass production measured in field trials as shoot length and dry mass and rust resistance measured by field screenings and in-vitro tests. DNA analysis of the willow rusts in stock collections and field trials revealed that S. daphnoides is exclusively infected by specialised fungus Melampsora epitea-typica f. sp. daphnoides. The selection process yielded 48 genotypes that have been planted in three comprehensive field trials. They serve as basis for further investigation in biomass, salicylate and phenolics production.
Dariusz Kruszka, Carolina Gomes, Andrea Pagano, Piotr Kachlicki, Jorge Paiva*
Institute of Plant Genetics of the Polish Academy of Sciences, 34 Strzeszynska street, 60-479 Poznan, Poland
*Corresponding author: jpai@igr.poznan.pl
Keywords: Purple willow, metabolomics, UPLC-HRMS, secondary metabolism
Willows (Salix spp.) are known as an economically important source of biomass for energy and biofuels, as tools for erosion control and phytoremediation, and for their use in medicine. For example, willow bark (Cortex Salicis) is an herbal raw material still used as an anti-inflammatory drug, as it contains salicin as a major phytochemical together with other bioactive compounds. Among the willow species, Salix pupurea (Purple willow) is being considered a good model species for bioenergy and phytochemical production, particularly since its genome has been made publicly available. Despite the importance of willows and the growing number of studies on these species, a systematic and comprehensive analysis of their metabolome is still lacking, as only few reports on willow metabolome are available in the current literature.
In the frame of the PurpleWalls project, untargeted metabolomic methodologies were applied to provide a comprehensive characterization of the metabolomic profiles of roots, internodes, xylem and bark of Salix purpurea. The tissue samples were collected from ramets of the ELB2/5genotype (Germplasm Collection of University of Warmia-Mazury) growing in greenhouse conditions for one or two months. Ultra-performance liquid chromatography coupled with high resolution mass spectrometry (UPLC-HRMS) was applied to profile the metabolites in the methanolic extracts from the different tissue samples. Additionally, GC-MS analysis was used to identify the metabolites in the cell wall fraction of the xylem samples. The mass spectrometry data were pre-processed using two chemoinformatic tools: MzMine and MSDIAL. Further analysis was performed in the MetaboAnalyst 4.0 software. The results from the analysed samples highlighted the accumulation of several phytochemical groups, such as hydroxybenzoic and hydroxycinnamic acid derivatives, several flavonoids (e.g.: naringenin, naringenin hexosides, astilbin, quercetin, procyanidins, catechins), lignans and coumarins. A large number of primary metabolites was also detected, including amino acids and different lipids. Moreover, several sugar compounds were identified using GC-MS. To our best knowledge, these results constitute the most comprehensive overview of the metabolites in Salix purpurea, and might contribute considerably to the enrichment of the Salix metabolite database (Alferis et al., 2015).
Acknowledgements:
This work was supported by the NCN project Sonata-bis UMO-2015/18/E/NZ2/00694 (PurpleWalls Project). JAPP acknowledges his research contract in the frame of EU-FP7-ERAChairs-PillotCAll-2013 project “Biotalent - The creation of the Department of Integrative Plant Biology” (FP7-REGPOT-621321) and the Polish financial sources for education in the years 2015–2019 allocated to an international co-financed project. We thanks Pawel Sulima and Jerzy Przyborowski (UWM, Poland) for made available the genotypes used in this analysis.
References: Aliferis et al. Front Plant Sci. 2015;6(MAY):1-15. doi:10.3389/fpls.2015.00344
Carolina Gomes1, Andrea Pagano1, Dariusz Kruska1, Pawel Sulima2, Jorge Almiro Pinto Paiva*
1Institute of Plant Genetic Polish Academy of Science, ul. Strzeszyńska 34, 60-479 Poznań, Poland
2Department of Plant Breeding and Seed Production, University of Warmia and Mazury in Olsztyn, Plac Łódzki 3, 10-724 Olsztyn, Poland
*Corresponding author: jpai@igr.poznan.pl
Fast growing woody plants such as Salix spp. represent an appealing source of renewable biomass feedstock for bio-energy. Secondary cell wall (SCW) chemical composition and structure have significant impacts on their uses as a source of raw material for different applications (timber, pulp, bioenergy). Therefore, increasing our knowledge of the molecular regulation of SCW biosynthesis and xylogenesis is a critical issue to develop and test new molecular-breeding strategies and innovative tailored-wood raw products for bio-energy production, for instance. PurpleWalls project aims at providing novel insights on the molecular regulatory mechanisms underlying secondary cell wall biosynthesis, with main focus on the transcription regulation (role of transcription factors, miRNAs, chromatin remodelling) and their impact on SCW properties, , using artificially altered purple willow (Salix purpurea) plants.
The main outputs of the PurpleWalls include:
* the development of a hairy root transformation protocol for Salix pupurea allowing the functional studies of key genes in this species,
* deep characterization of the DNA methylation landscape by using WGBS-seq andd of the transcriptome by RNA-seq (including ncRNAs), and deep characterization of metabolome, and
* Identification of S purpurea DNA methylation toolbox
Moreover, the project offered training opportunities to 14 national and international students in collaboration with UAM and PULS (Poznan, Poland) and University of Pavia (Italy).
Acknowledgments: NCN (Poland) under the project SONATA Bis 5 “PurpleWalls” (UMO-2015/18/E/NZ2/00694). JAPP thanks the his research contract EU FP7 BIOTALENT project [GA621321] and financial sources for education in the years 2015-2019 allocated to an international co-financed project". Special acknowledgement to Dr. Pawel Sulima and Prof. Jerzy Przyborowski (Warmia-Mazuria University, in Olsztyn, Poland) for the access to Salix purpurea germplasm collection, Jacqueline Grima-Pettenati for the opportunity of developing the hairy roots transformation system, and the consultors, Susana Araújo, Piotr Kachlicki, and Stephane Maury for the precious advises given along project.
Karolina Ratajczak1, Katarzyna Panasiewicz1, Alicja Niewiadomska2, Hanna Sulewska1, Agnieszka Wolna-Maruwka2, Klaudia Borowiak3, Anna Budka4
1Department of Agronomy, Poznań University of Life Sciences, Poznań, Poland
2Department of General and Environmental Microbiology, Poznań University of Life Sciences, Poznań, Poland
3Department of Ecology and Environmental and Protection, Poznań University of Life Sciences, Poznań, Poland
4Department of Mathematical and Statistical Methods, Poznań University of Life Sciences, Poznań, Poland
Key words: photosynthesis activity, willow, yields, bacteria, sewage sludge
Willow (Salix viminalis L.) is not only a good source of biomass, but also a good phytoremediator, that is capable of removing hazardous substances from the contaminated environment. Currently it is necessary to find appropriate methods of sewage sludge management, which constitutes a considerable ecological and economic problem, because it is predicted that the total amount of sewage sludge produced in the EU may reach 13 million tons in 2020. The optimal solution is to use sludge as a fertiliser for industrial plants, including energy crops, like willow that is not used in food production. Thus the aim of this study was to assess the effect of application of sewage sludge, the bacteria, actinobacteria, fungi microbial inoculant (BAF), sewage sludge + BAF on growth, photosynthesis activity and yield of willow as an energy crop, as well as the biochemical and microbial activity of soil in such energy crop willow culture. Experiments were conducted in the years 2013–2015 on experimental fields of the Poznan University of Life Sciences. The results indicate that the application of sewage sludge increased yields and improved selected photosynthesis activity and biometric traits of willow.
The help with the field trials and measurements provided by the the Research and Education Center Gorzyń, branch in Złotniki is kindly acknowledged.