Comparative de novo transcriptome analysis of barley varieties with different malting qualities.

Barley is one of the most important crops in the world. Barley is used as both food and feed and is important for malt production. Demands for malting quality differ among countries and customs. Malting quality is a complex characteristic involving barley genetics, the environmental conditions during barley growth, and the technological parameters of the malting process. In this study, the hypothesis was that there were no differences between two groups of barley varieties with different but defined malting qualities, which was tested using RNA sequencing during selected stages of malting. In total, 919 differentially transcribed genes between the two barley groups were identified and annotated. Differentially expressed genes (DEGs) were primarily assigned to gene ontology (GO) terms of oxidation-reduction process - oxidoreductase activity, response to stress, carbohydrate metabolic process, and proteolysis - hydrolase activity, and metal ion binding. Genes connected with the plasma membrane and its integral components also play important roles in malting quality. DEG profiles of selected genes in the three malting stages indicate a complex character of malting quality. Many single-nucleotide polymorphisms (SNPs) and insertions and deletions (indels) were identified. SNPs and indels with the best quality were used for primer design. After optimization and validation, five molecular markers were developed for use in barley breeding.

Oats as a Safe Alternative to Triticeae Cereals for People Suffering from Celiac Disease? A Review.

Oats represent a promising alternative to small-grain cereals from Triticeae group (wheat, barley, rye) for persons suffering from any form of gluten intolerance, especially celiac disease (CD), since oat-specific prolamins avenins reveal generally lower gluten content and immunoreactivity. Recent studies on avenin molecular structure revealed large genetic variability in avenin sequences affecting the spectrum of gluten peptides produced by hydrolases in human digestive tract. The aim of the present review is to summarise recent knowledge obtained in laboratory studies focused on the effect of avenin-derived peptides on reactivity of crucial components of human immune system such as dendritic cells (DC) and T-cells. The other part of the review summarises the results of clinical studies with CD patients including oat products in their diet. Since different clinical studies revealed contradictory results regarding potential safety of oats for CD patients, the focus has to be directed at genetic variability in oat avenins. Identification of avenin isoforms with minimum CD immunoreactivity will open up ways leading to designing novel oat cultivars suitable for CD patients. Knowledge on immunoreactivity of gluten peptides together with breeding new oat cultivars revealing minimum avenin immunoreactivity with respect to CD as well as application of food processing technologies leading to gluten content reduction should result in development of gluten-free oats safe for celiacs.

Changes in polyphenol compounds and barley laccase expression during the malting process.

BACKGROUND: Polyphenols and phenolic acid are able to slow down or prevent oxidation processes and are therefore thought to have important effects in malting and brewing. Laccase catalyses the oxidation of a wide variety of substrates, including polyphenols. The aim of this paper was to determine the changes in polyphenol compounds and the relative expression of the HvLac1 gene during malting. RESULTS: The dominant phenolic acid was ferulic acid. The amount of ferulic acid increased, whereas the amount of vanillic acid decreased during malting. The highest levels of expression of the HvLac1 gene were observed during the third air rest period in varieties with the 'Haruna Nijo' (HN) allele, as recommended for the production of beer with the protected geographical indication (PGI) 'Ceske pivo' (Czech beer), whereas the highest expression was observed in the first day of germination in varieties with the 'Morex' (M) allele. However, the profiles of HvLac1 gene expression in varieties with alternative alleles during malting were similar, and the level of polyphenol compounds throughout malting was different. CONCLUSION: The polyphenol contents in barley increased several-fold during malting, and the degree of increase differed with variety. The expression of HvLac1 transcript was similar in every barley variety.

Analysis of oat seed transcriptome with regards to proteins involved in celiac disease.

Oat (Avena sativa L.) is considered to be a healthy food. In contrast to other grain crops, oat is high in protein, lipids, dietary fiber, antioxidants, and uniquely in avenanthramides. The question of whether it can also be consumed by people suffering from celiac disease is still unresolved. The main aim of this study was to extract and sequence genes for potentially harmful avenins, globulins, and α-amylase/trypsin inhibitors in six oat varieties and to establish their variability using PacBio sequencing technology of enriched libraries. The results were compared with sequences of the genes already present in databases. In total, 21 avenin, 75 globulin, and 25 α-amylase/trypsin inhibitor genes were identified and mapped in the hexaploid oat chromosomes. In all of the three gene families, only marginal sequence differences were found between the oat varieties within the individual genes. Avenin epitopes were found in all four types of avenin genes occurring in all oat varieties tested within this study. However, the number of avenin genes was nearly four times lower than of globulin genes and, on the protein level, formed only 10% of storage proteins. Therefore, the question of whether oat is safe to celiac disease people is a question of boundary values.

Molecular Characterization of Novel x-Type HMW Glutenin Subunit 1B × 6.5 in Wheat.

A novel high molecular weight glutenin subunit encoded by the Glu-1B locus was identified in the French genotype Bagou, which we named 1B × 6.5. This subunit differed in SDS-PAGE from well-known 1B × 6 and 1B × 7 subunits, which are also encoded at this locus. Subunit 1B × 6.5 has a theoretical molecular weight of 88,322.83 Da, which is more mobile than 1B × 6 subunit, and isoelectric point (pI) of about 8.7, which is lower than that for 1B × 6 subunit. The specific primers were designed to amplify and sequence 2476 bp of the Glu-1B locus from genotype Bagou. A high level of similarity was found between the sequence encoding 1B × 6.5 and other x-type encoding alleles of this locus.

Efficient Confirmation of Plant Viral Proteins and Identification of Specific Viral Strains by nanoLC-ESI-Q-TOF Using Single-Leaf-Tissue Samples.

Plant viruses are important pathogens that cause significant crop losses. A plant protein extraction protocol that combines crushing the tissue by a pestle in liquid nitrogen with subsequent crushing by a roller-ball crusher in urea solution, followed by RuBisCO depletion, reduction, alkylation, protein digestion, and ZipTip purification allowed us to substantially simplify the sample preparation by removing any other precipitation steps and to detect viral proteins from samples, even with less than 0.2 g of leaf tissue, by a medium resolution nanoLC-ESI-Q-TOF. The presence of capsid proteins or polyproteins of fourteen important viruses from seven different families (Geminiviridae, Luteoviridae, Bromoviridae, Caulimoviridae, Virgaviridae, Potyviridae, and Secoviridae) isolated from ten different economically important plant hosts was confirmed through many identified pathogen-specific peptides from a protein database of host proteins and potential pathogen proteins assembled separately for each host and based on existing online plant virus pathogen databases. The presented extraction protocol, combined with a medium resolution LC-MS/MS, represents a cost-efficient virus protein confirmation method that proved to be effective at identifying virus strains (as demonstrated for PPV, WDV) and distinct disease species of BYDV, as well as putative new viral protein sequences from single-plant-leaf tissue samples. Data are available via ProteomeXchange with identifier PXD022456.

Genetic differentiation of Rubus chamaemorus populations in the Czech Republic and Norway after the last glacial period.

The population structure of cloudberry (Rubus chamaemorus L.), collected from Krkonose Mountains (the Czech Republic), continental Norway and Spitsbergen, was examined using microsatellite analyses (SSR). Among 184 individuals, 162 different genotypes were identified. The overall unbiased gene diversity was high ( h^=0.463 ). A high level of genetic differentiation among populations (FST = 0.45; p < .01) indicated restricted gene flow between populations. Using a Bayesian approach, six clusters were found which represented the genetic structure of the studied cloudberry populations. The value of correlation index between genetic and geographical distances (r = .44) indicates that gene flow, even over a long distance, could exist. An exact test of population differentiation showed that Rubus chamaemorus populations from regions (Krkonose Mountains, continental Norway and Spitsbergen) are differentiated although some individuals within populations share common alleles even among regions. These results were confirmed by AMOVA, where the highest level of diversity was found within populations (70.8%). There was no difference between 87 pairs of populations (18.7%) mostly within cloudberry populations from continental Norway and from Spitsbergen. Based on obtained results, it is possible to conclude that Czech and Norwegian cloudberry populations are undergoing differentiation, which preserves unique allele compositions most likely from original populations during the last glaciation period. This knowledge will be important for the creation and continuation of in situ and ex situ conservation of cloudberry populations within these areas.

Genetic structure of Pyrenophora teres net and spot populations as revealed by microsatellite analysis.

The population structure of the fungal pathogen Pyrenophora teres, collected mainly from different regions of the Czech and Slovak Republics, was examined using a microsatellite analyses (SSR). Among 305 P. teres f. teres (PTT) and 82 P. teres f. maculata (PTM) isolates that were collected, the overall gene diversity was similar (h = 0.12 and h = 0.13, respectively). A high level of genetic differentiation (FST = 0.46; P < 0.001) indicated the existence of population structure. Nine clusters that were found using a Bayesian approach represent the genetic structure of the studied P. teres populations. Two clusters consisted of PTM populations; PTT populations formed another seven clusters. An exact test of population differentiation confirmed the results that were generated by Structure. There was no difference between naturally infected populations over time, and genetic distance did not correlate with geographical distance. The facts that all individuals had unique multilocus genotypes and that the hypothesis of random mating could not be rejected in several populations or subpopulations serve as evidence that a mixed mating system plays a role in the P. teres life cycle. Despite the fact that the genetic differentiation value between PTT and PTM (FST = 0.30; P < 0.001) is lower than it is between the populations within each form (FST = 0.40 (PTT); FST = 0.35 (PTM); P < 0.001) and that individuals with mixed PTT and PTM genomes were found, the two forms of P. teres form genetically separate populations. Therefore, it can be assumed that these populations have most likely undergone speciation.