Project description:Flax (Linum usitatissimum L.) is a multipurpose crop which is used for the production of textile, oils, composite materials, pharmaceuticals, etc. Soil acidity results in a loss of seed and fiber production of flax, and aluminum toxicity is a major factor that depresses plant growth and development in acid conditions. In the present work, we evaluated gene expression alterations in four flax genotypes with diverse tolerance to aluminum exposure. Using RNA-Seq approach, we revealed genes that are differentially expressed under aluminum stress in resistant (Hermes, TMP1919) and sensitive (Lira, Orshanskiy) cultivars and selectively confirmed the identified alterations using qPCR. To search for differences in response to aluminum between resistant and sensitive genotypes, we developed the scoring that allowed us to suggest the involvement of MADS-box and NAC transcription factors regulating plant growth and development and enzymes participating in cell wall modifications in aluminum tolerance in flax. Using Gene Ontology (GO) enrichment analysis, we revealed that glutathione metabolism, oxidoreductase, and transmembrane transporter activities are the most affected by the studied stress in flax. Thus, we identified genes that are involved in aluminum response in resistant and sensitive genotypes and suggested genes that contribute to flax tolerance to the aluminum stress.
Project description:Flax phloem fibers achieve their length by intrusive-diffusive growth, which requires them to penetrate the extracellular matrix of adjacent cells. Fiber elongation therefore involves extensive remodelling of cell walls and middle lamellae, including modifying the degree and pattern of methylesterification of galacturonic acid (GalA) residues of pectin. Pectin methylesterases (PME) are important enzymes for fiber elongation as they mediate the demethylesterification of GalA in muro, in either a block-wise fashion or in a random fashion. Our objective was to identify PMEs and PMEIs that mediate phloem fiber elongation in flax. For this purpose, we measured transcript abundance of candidate genes at nine different stages of stem and fiber development and found sets of genes enriched during fiber elongation and maturation as well as during xylem development. We expressed one of the flax PMEIs in E. coli and demonstrated that it was able to inhibit most of the native PME activity in the upper portion of the flax stem. These results identify key genetic components of the intrusive growth process and define targets for fiber engineering and crop improvement.
Project description:Flaxseed oil is characterized by high content of essential polyunsaturated fatty acids (PUFA) promoted as a human dietary supplement protecting against atherosclerosis. The disadvantage of the high PUFA content in flax oil is high susceptibility to oxidation, which can result in carcinogenic compound formation. Linola flax cultivar is characterized by high linoleic acid content in comparison to traditional flax cultivars rich in linolenic acid. The changes in fatty acid proportions increase oxidative stability of Linola oil and broaden its use as an edible oil for cooking. However one of investigated transgenic lines has high ALA content making it suitable as omega-3 source. Protection of PUFA oxidation is a critical factor in oil quality. The aim of this study was to investigate the impact of phenylpropanoid contents on the oil properties important during the whole technological process from seed storage to grinding and oil pressing, which may influence health benefits as well as shelf-life, and to establish guidelines for the selection of new cultivars.The composition of oils was determined by chromatographic (GS-FID and LC-PDA-MS) methods. Antioxidant properties of secondary metabolites were analyzed by DPPH method. The stability of oils was investigated: a) during regular storage by measuring acid value peroxide value p-anisidine value malondialdehyde, conjugated dienes and trienes; b) by using accelerated rancidity tests by TBARS reaction; c) by thermoanalytical - differential scanning calorimetry (DSC).In one approach, in order to increase oil stability, exogenous substances added are mainly lipid soluble antioxidants from the isoprenoid pathway, such as tocopherol and carotene. The other approach is based on transgenic plant generation that accumulates water soluble compounds. Increased accumulation of phenolic compounds in flax seeds was achieved by three different strategies that modify genes coding for enzymes from the phenylpropanoid pathway. The three types of transgenic flax had different phenylpropanoid profiles detected in oil, highly increasing its stability.We found that hydrophilic phenylpropanoids more than lipophilic isoprenoid compounds determine oil stability however they can work synergistically. Among phenolics the caffeic acid was most effective in increasing oil stability.
Project description:A robust phenotypic plasticity to ward off adverse environmental conditions determines performance and productivity in crop plants. Flax (linseed), is an important cash crop produced for natural textile fiber (linen) or oilseed with many health promoting products. This crop is prone to drought stress and yield losses in many parts of the world. Despite recent advances in drought research in a number of important crops, related progress in flax is very limited. Since, response of this plant to drought stress has not been addressed at the molecular level; we conducted microarray analysis to capture transcriptome associated with induced drought in flax. This study identified 183 differentially expressed genes (DEGs) associated with diverse cellular, biophysical and metabolic programs in flax. The analysis also revealed especially the altered regulation of cellular and metabolic pathways governing photosynthesis. Additionally, comparative transcriptome analysis identified a plethora of genes that displayed differential regulation both spatially and temporally. These results revealed co-regulated expression of 26 genes in both shoot and root tissues with implications for drought stress response. Furthermore, the data also showed that more genes are upregulated in roots compared to shoots, suggesting that roots may play important and additional roles in response to drought in flax. With prolonged drought treatment, the number of DEGs increased in both tissue types. Differential expression of selected genes was confirmed by qRT-PCR, thus supporting the suggested functional association of these intrinsic genes in maintaining growth and homeostasis in response to imminent drought stress in flax. Together the present study has developed foundational and new transcriptome data sets for drought stress in flax.
Project description:We used RNAseq to compare transcript expression patterns in two segments of the vegetative stem of 14d flax plants, from which all visible leaves had been removed. The segments were: (i) the apical region (AR) of the shoot apex, which contained the apical-most 0.5 mm of the stem, including the SAM and its immediate derivatives; and (ii) the basal region (BR), which contained the entire stem except for the apical-most 1 cm, and therefore represented all stem and vascular tissues at later stages of differentiation as compared to the AR. These data will help identify genes that contribute to specification of phloem fiber identity.
Project description:Three 2cm segments were excised from different parts (TOP, MID, BOT) along the vertical axis of a 4 week old (25cm) stem of flax (L. usitatissimum) were compared using a cDNA amplicon array. Each segment represented a different developmental stage, especially in relation to bast fibre differentiation (i.e. TOP= elongation, MID=transition, BOT= thickening). Only the cDNAs that showed the highest differential expression were sequenced.
Project description:New flaxseed cultivars differing in seed mucilage content (MC) with low hull content (HC) represent an attractive option to simultaneously target the food and feed markets. Here, a genome-wide association study (GWAS) was conducted for MC and HC in 200 diverse flaxseed accessions genotyped with 1.7 million single nucleotide polymorphism (SNP) markers. The data obtained for MC and HC indicated a broad phenotypic variation and high (~70%) and a moderate (~49%) narrow sense heritability, respectively. MC and HC did not differ statistically between fiber and oil morphotypes, but yellow-seeded accessions had 2.7% less HC than brown-seeded ones. The genome-wide linkage disequilibrium (LD) decayed to r² = 0.1 at a physical distance of ~100 kb. Seven and four quantitative trait loci (QTL) were identified for MC and HC, respectively. Promising candidate genes identified include Linum usitatissimum orthologs of the Arabidopsis thaliana genes TRANSPARENT TESTA 8, SUBTILISIN-LIKE SERINE PROTEASE, GALACTUROSYL TRANSFERASE-LIKE 5, MUCILAGE-MODIFIED 4, AGAMOUS-LIKE MADS-BOX PROTEIN AGL62, GLYCOSYL HYDROLASE FAMILY 17, and UDP-GLUCOSE FLAVONOL 3-O-GLUCOSYLTRANSFERASE. These genes have been shown to play a role in mucilage synthesis and release, seed coat development and anthocyanin biosynthesis in A. thaliana. The favorable alleles will be useful in flaxseed breeding towards the goal of achieving the ideal MC and HC composition for food and feed by genomic-based breeding.
Project description:Membrane intrinsic proteins (MIPs) form transmembrane channels and facilitate transport of myriad substrates across the cell membrane in many organisms. Majority of plant MIPs have water transporting ability and are commonly referred as aquaporins (AQPs). In the present study, we identified aquaporin coding genes in flax by genome-wide analysis, their structure, function and expression pattern by pan-genome exploration. Cross-genera phylogenetic analysis with known aquaporins from rice, arabidopsis, and poplar showed five subgroups of flax aquaporins representing 16 plasma membrane intrinsic proteins (PIPs), 17 tonoplast intrinsic proteins (TIPs), 13 NOD26-like intrinsic proteins (NIPs), 2 small basic intrinsic proteins (SIPs), and 3 uncharacterized intrinsic proteins (XIPs). Amongst aquaporins, PIPs contained hydrophilic aromatic arginine (ar/R) selective filter but TIP, NIP, SIP and XIP subfamilies mostly contained hydrophobic ar/R selective filter. Analysis of RNA-seq and microarray data revealed high expression of PIPs in multiple tissues, low expression of NIPs, and seed specific expression of TIP3 in flax. Exploration of aquaporin homologs in three closely related Linum species bienne, grandiflorum and leonii revealed presence of 49, 39 and 19 AQPs, respectively. The genome-wide identification of aquaporins, first in flax, provides insight to elucidate their physiological and developmental roles in flax.
Project description:BackgroundFlax (Linum usitatissimum L.) is an important crop for the production of bioproducts derived from its seed and stem fiber. Transposable elements (TEs) are widespread in plant genomes and are a key component of their evolution. The availability of a genome assembly of flax (Linum usitatissimum) affords new opportunities to explore the diversity of TEs and their relationship to genes and gene expression.ResultsFour de novo repeat identification algorithms (PILER, RepeatScout, LTR_finder and LTR_STRUC) were applied to the flax genome assembly. The resulting library of flax repeats was combined with the RepBase Viridiplantae division and used with RepeatMasker to identify TEs coverage in the genome. LTR retrotransposons were the most abundant TEs (17.2% genome coverage), followed by Long Interspersed Nuclear Element (LINE) retrotransposons (2.10%) and Mutator DNA transposons (1.99%). Comparison of putative flax TEs to flax transcript databases indicated that TEs are not highly expressed in flax. However, the presence of recent insertions, defined by 100% intra-element LTR similarity, provided evidence for recent TE activity. Spatial analysis showed TE-rich regions, gene-rich regions as well as regions with similar genes and TE density. Monte Carlo simulations for the 71 largest scaffolds (? 1 Mb each) did not show any regional differences in the frequency of TE overlap with gene coding sequences. However, differences between TE superfamilies were found in their proximity to genes. Genes within TE-rich regions also appeared to have lower transcript expression, based on EST abundance. When LTR elements were compared, Copia showed more diversity, recent insertions and conserved domains than the Gypsy, demonstrating their importance in genome evolution.ConclusionsThe calculated 23.06% TE coverage of the flax WGS assembly is at the low end of the range of TE coverages reported in other eudicots, although this estimate does not include TEs likely found in unassembled repetitive regions of the genome. Since enrichment for TEs in genomic regions was associated with reduced expression of neighbouring genes, and many members of the Copia LTR superfamily are inserted close to coding regions, we suggest Copia elements have a greater influence on recent flax genome evolution while Gypsy elements have become residual and highly mutated.
Project description:Pasmo is one of the most widespread diseases threatening flax production. To identify genetic regions associated with pasmo resistance (PR), a genome-wide association study was performed on 370 accessions from the flax core collection. Evaluation of pasmo severity was performed in the field from 2012 to 2016 in Morden, MB, Canada. Genotyping-by-sequencing has identified 258,873 single nucleotide polymorphisms (SNPs) distributed on all 15 flax chromosomes. Marker-trait associations were identified using ten different statistical models. A total of 692 unique quantitative trait nucleotides (QTNs) associated with 500 putative quantitative trait loci (QTL) were detected from six phenotypic PR datasets (five individual years and average across years). Different QTNs were identified with various statistical models and from individual PR datasets, indicative of the complementation between analytical methods and/or genotype × environment interactions of the QTL effects. The single-locus models tended to identify large-effect QTNs while the multi-loci models were able to detect QTNs with smaller effects. Among the putative QTL, 67 had large effects (3-23%), were stable across all datasets and explained 32-64% of the total variation for PR in the various datasets. Forty-five of these QTL spanned 85 resistance gene analogs including a large toll interleukin receptor, nucleotide-binding site, leucine-rich repeat (TNL) type gene cluster on chromosome 8. The number of QTL with positive-effect or favorite alleles (NPQTL) in accessions was significantly correlated with PR (R 2 = 0.55), suggesting that these QTL effects are mainly additive. NPQTL was also significantly associated with morphotype (R 2 = 0.52) and major QTL with positive effect alleles were present in the fiber type accessions. The 67 large effect QTL are suited for marker-assisted selection and the 500 QTL for effective genomic prediction in PR molecular breeding.