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Background: Wood represents the majority of the biomass on land and constitutes a renewable source of biofuels and other bioproducts. However, wood is recalcitrant to bioconversion, raising a need for feedstock improvement in production of, for instance, biofuels. We investigated the properties of wood that affect bioconversion, as well as the underlying genetics, to help identify superior tree feedstocks for biorefining.

Results: We recorded 65 wood-related and growth traits in a population of 113 natural aspen genotypes from Sweden (https://doi.org/10.5061/dryad.gtht76hrd). These traits included three growth and field performance traits, 20 traits for wood chemical composition, 17 traits for wood anatomy and structure, and 25 wood saccharification traits as indicators of bioconversion potential. Glucose release after saccharification with acidic pretreatment correlated positively with tree stem height and diameter and the carbohydrate content of the wood, and negatively with the content of lignin and the hemicellulose sugar units. Most of these traits displayed extensive natural variation within the aspen population and high broad-sense heritability, supporting their potential in genetic improvement of feedstocks towards improved bioconversion. Finally, a genome-wide association study (GWAS) revealed 13 genetic loci for saccharification yield (on a whole-tree-biomass basis), with six of them intersecting with associations for either height or stem diameter of the trees.

Conclusions: The simple growth traits of stem height and diameter were identified as good predictors of wood saccharification yield in aspen trees. GWAS elucidated the underlying genetics, revealing putative genetic markers for bioconversion of bioenergy tree feedstocks.

• The genetic control underlying natural variation in lignin content and composition in trees is not fully understood. We performed a systems genetic analysis to uncover the genetic regulation of lignin biosynthesis in a natural ‘SwAsp’ population of aspen (Populus tremula) trees.
• We analyzed gene expression by RNA sequencing (RNA-seq) in differentiating xylem tissues, and lignin content and composition using Pyrolysis-GC-MS in mature wood of 268 trees from 99 genotypes.
• Abundant variation was observed for lignin content and composition, and genome-wide association study identified proteins in the pentose phosphate pathway and arabinogalactan protein glycosylation among the top-ranked genes that are associated with these traits. Variation in gene expression and the associated genetic polymorphism was revealed through the identification of 312 705 local and 292 003 distant expression quantitative trait loci (eQTL). A co-expression network analysis suggested modularization of lignin biosynthesis and novel functions for the lignin-biosynthetic CINNAMYL ALCOHOL DEHYDROGENASE 2 and CAFFEOYL-CoA O-METHYLTRANSFERASE 3. PHENYLALANINE AMMONIA LYASE 3 was co-expressed with HOMEOBOX PROTEIN 5 (HB5), and the role of HB5 in stimulating lignification was demonstrated in transgenic trees.
• The systems genetic approach allowed linking natural variation in lignin biosynthesis to trees´ responses to external cues such as mechanical stimulus and nutrient availability.

Wood decaying fungi cause substantial economic losses due to discoloration and loss of biomass. The wood of European aspen, Populus tremula (L.), is normally very light and uniform in color, but it is prone to fungal infections that are poorly described. We observed discoloration in woody tissues of five-year-old aspen ramets that were formed on stools of trees cut in the year 2014. The trees were part of the Swedish aspen (SwAsp) collection, which comprises 112 genotypes grown in a common garden experiment established in 2004 in Ekebo, southern Sweden. Internal transcribed spacer sequencing (ITS2) of the fungal rRNA genes in 19 SwAsp trees with visible symptoms of fungal infection revealed Cadophora and Ascocoryne as the dominating fungal genera in the susceptible trees. These fungi have not previously been associated with aspen wood discoloration or decay. Chemical analysis by pyrolysis-GC/MS revealed a positive correlation between the abundance of the fungal infections (wood discoloration) and the relative abundance of p-hydroxyphenyl (H) type lignin. Molecular mechanisms underlying the pathogenesis were investigated with a genome-wide association study (GWAS) of the fungal discoloration area in the SwAsp collection, and an existing RNA-sequencing dataset from the same material. One genomic locus and several genes with putative roles in pathogenesis or wood formation were linked to symptoms related to the fungal infections. These loci provide potential genetic markers that could be used to trace the abundance of the fungal pathogens in young aspen stems or to breed trees towards increased fungal resistance.