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Wheat is one of the world’s crucial staple food crops. In turn, einkorn wheat (Triticum monococcum L.) is considered a wild relative of wheat (Triticum aestivum L.) and can be used as a source of agronomically important genes for breeding purposes. Cultivated T. monococcum subsp. monococcum originated from T. monococcum subsp. aegilopoides (syn. T. boeticum). For the better utilization of valuable genes from these species, it is crucial to discern the genetic diversity at their cytological and molecular levels. Here, we used a fluorescence in situ hybridization toolbox and molecular markers linked to the leaf rust resistance gene Lr63 (located on the short arm of the 3Am chromosome—3AmS) to track the polymorphisms between T. monococcum subsp. monococcum, T. boeticum and T. urartu (A-genome donor for hexaploid wheat) accessions, which were collected in different regions of Europe, Asia, and Africa. We distinguished three groups of accessions based on polymorphisms of cytomolecular and leaf rust resistance gene Lr63 markers. We observed that the cultivated forms of T. monococcum revealed additional marker signals, which are characteristic for genomic alternations induced by the domestication process. Based on the structural analysis of the 3AmS chromosome arm, we concluded that the polymorphisms were induced by geographical dispersion and could be related to adaptation to local environmental conditions.

Triticale (×Triticosecale Wittmack) is a commercial hybrid harboring wheat (Triticum sp.) and rye (Secale cereale L.) genomes. The limited genetic diversity of this crop resulted in the collapse of fungal disease resistance. Leaf rust disease, caused by Puccinia triticina Eriks. is reported to reduce the triticale yield significantly (more than 30%). There is a need to enlarge the genetic variability of this crop including leaf resistance genes. The main aim of this research was to transfer Lr39 and Lr54 leaf rust resistance genes into triticale from Aegilops tauschii and Ae. kotschyi, respectively. A reaction of seedlings of 200 plants of two triticale-Aegilops translocation lines (Bogo-2Dt.2R and Sekundo-2Sk.2R) was compared after inoculation with a natural mixture of P. triticina races, specific to triticale. Before inoculation, each plant was screened using molecular cytogenetics and molecular markers linked to leaf rust resistance genes. Presence of Aegilops chromosome segments was confirmed using genomic in situ hybridization (GISH). Lr39 and Lr54 leaf rust resistance genes were identified using Xgdm35 and S14 molecular markers, respectively. After inoculation, a significant improvement of resistance severity was observed in Sekundo-2Sk.2R in comparison with triticale cv. Sekundo plants. The resistance level of Bogo-2Dt.2R did not differ compared to triticale cv. Bogo plants. It was shown, that Lr39 gene did not increase the leaf rust resistance level of triticale cv. Bogo.

Currently, production of wheat cultivars (Triticum aestivum L.) that show durable field resistance against fungal pathogens is a priority of many breeding programs. This type of resistance involves race-nonspecific mechanisms and can be identified at adult-plant stages. Until now, seven genes (Lr34/Yr18, Lr46/Yr29, Lr67/Yr46, Lr68, Lr75, Lr77 and Lr78) conferring durable types of resistance against multiple fungal pathogens have been identified in the wheat gene pool. In this study we showed a multiplex Polymerase Chain Reaction (multiplex PCR) assay, which was developed for detection of slow rusting resistance genes Lr34, Lr46, Lr68, using molecular markers: csLV34, Xwmc44 and csGS, respectively. Identification of molecular markers was performed on 40 selected wheat genotypes which are the sources of slow rusting genes according to literature reports. Multiplex PCR is an important tool to reduce the time and cost of analysis. This multiplex PCR protocol can be applicable for genotyping processes and marker assisted resistance breeding of wheat.

Lr46/Yr29/Pm39 (Lr46) is a gene for slow rusting resistance in wheat. The aim of the study was to analyze the miRNA expression in selected common wheat cultivars carrying resistance genes, Lr46 among others (HN Rod, Pavon‘S’, Myna‘S’, Frontana‘S’, and Sparrow’S’) in response to leaf rust infection caused by Puccinia triticina Erikss. In the Pavon ‘S’, Myna ‘S’, Frontana‘S’, and Sparow‘S’ varieties a product with a length of 242 bp has been identified, which is specific to the Xwmc44 marker linked to the brown rust resistance gene Lr46. In the next step, the differences in the expression of microRNA (miR5085 and miR164) associated with the Lr46 gene, which is responsible for different resistance of selected wheat cultivars to leaf rust, were examined using emulsion PCR (ddPCR). In the experiment, biotic stress was induced in mature plants by infecting them with fungal spores under controlled conditions in a growth chamber. For analysis the plant material was collected before inoculation and 6, 12, 24, and 48 h after inoculation. The experiments also showed that plant infection with Puccinia triticina resulted in an increase in miR164 expression in cultivars carrying the Lr46 gene. The expression of miR164 remained stable in a control cultivar (HN ROD) lacking this gene. This has proved that miR164 can be involved in leaf rust resistance mechanisms.

Recently, leaf rust and yellow rust caused by the fungi Puccinia triticina Erikss. and P. striiformis Westend f. sp. tritici Eriks and Henn are diseases of increasing threat in triticale (× Triticosecale Wittmack, AABBRR, 2n = 6x = 42) growing areas. The use of genetic resistance is considered the most economical, effective and environmentally friendly method to control the disease and minimize the use of fungicides. Currently, breeding programs mainly relied on race-specific Lr and Yr genes (R), but new races of the rust fungi frequently defeat resistance. There is a small group of genes that causes partial type of resistance (PR) that are characterized by a slow epidemic build up despite a high infection type. In wheat slow rusting resistance genes displayed longer latent periods, low infection frequencies, smaller pustule size and less spore production. Slow rusting Lr46/Yr29 gene, located on chromosome 1B, is being exploited in many wheat breeding programs. So far, there is no information about slow rusting genes in triticale. This paper showed significant differences between the results of identification of wheat molecular markers Xwmc44 and csLV46G22 associated with Lr46/Yr29 in twenty triticale cultivars, which were characterized by high levels of field resistance to leaf and yellow rust. The csLV46G22res marker has been identified in the following cultivars: Kasyno, Mamut and Puzon. Belcanto and Kasyno showed the highest resistance levels in three-year (2016–2018), leaf and yellow rust severity tests under post-registration variety testing program (PDO). Leaf tip necrosis, a phenotypic trait associated with Lr34/Yr18 and Lr46/Yr29 was observed, among others, to Belcanto and Kasyno, which showed the highest resistance for leaf rust and yellow rust. Kasyno could be considered to have Lr46/Yr29 and can be used as a source of slow rust resistance in breeding and importantly as a component of gene pyramiding in triticale.

A number of studies have shown that the greater the genetic diversity of parental lines, the greater the heterosis effect. Genetic or phenotypic variation can be estimated by genotype testing on the basis of the observations obtained through prediction (a priori) or the observations and studies (a posteriori). The first method uses data such as the genealogy of a given subject and the information about its geographical origin. The second method is based on the phenotypic observation and studies, as well as on the molecular research. The development of molecular genetics and genotype testing methods at the DNA level has made it possible to rapidly assess the genetic variability regardless of the modifying effect of the environment. The aim of this study was to determine the relationship between the degree of relatedness and the DNA polymorphism (determined using AFLP, RAPD, and SSR markers) of inbred maize lines and the effect of hybrid-form heterosis. Our analysis demonstrated that the parental components for heterosis crosses can be selected on the basis of the genetic similarity determined using the molecular SSR markers and the Jaccard, Kluczyński, Nei, and Rogers coefficients. Molecular AFLP markers proved less useful for selecting the parental components, but may be used to group lines with incomplete origin data. In the case of the RAPD markers, no clear relationship between genetic distance and the heterosis effect was found in this study.

Alien chromosome introgression has become a valuable tool to broaden the genetic variability of crop plants via chromosome engineering. This study details the procedure to obtain monosomic addition and monosomic substitution lines of the triticale carrying 2Skchromosome fromAegilops kotchyiBoiss., which harborsLr54+Yr37leaf and stripe rust-resistant gene loci, respectively. Initially,A. kotschyi×Secale cerealeartificial amphiploids (2n= 6x= 42 chromosomes, UUSSRR) were crossed with triticale cv. “Sekundo” (2n= 6x= 42, AABBRR) in order to obtain fertile offspring. Cyto-molecular analyses of five subsequent backcrossing generations revealed that 2Skchromosome was preferentially transmitted. This allowed for the selection of monosomic 2Skaddition (MA2Sk) lines of triticale. Finally, the 2Sk(2R) substitution plants were obtained by crossing MA2Skwith the nullisomic (N2R) plants of triticale. The presence of 2Skchromosome in subsequent generations of plants was evaluated using SSR markers linked toLr54+Yr37loci. Disease evaluation of the monosomic 2Sk(2R) substitution plants for the reaction to leaf and stripe rust infection were carried out under controlled conditions in a growth chamber. The results showed significant improvement of leaf rust resistance severity of monosomic substitution plants compared with control (“Sekundo”). In contrast, the introgression of theLr54+Yr37loci did not lead to improvement of stripe rust resistance. In summary, the creation of monosomic addition and monosomic substitution lines of triticale is the starting point for the precise and guided transfer ofLr54+Yr37loci. The results showed that the developed materials could be exploited for the development of triticale varieties with resistance to leaf rust.

There is a growing interest in breeding and production of hexaploid triticale (× Triticosecale Wittmack ex A. Camus) in European Union and in the world. It is reported that triticale can be an alternative to wheat (Triticum aestivum L.) for livestock feed production and has a potential to become preferred industrial energy crop. Fungal diseases, mainly leaf and stripe rusts, are the limiting factors of triticale growth and yield. Geneticists and breeders are now focusing on accumulation of the major genes for durability of rust resistance. Slow-rusting genes Lr34/Yr18 and Lr46/Yr19 are being exploited in many wheat breeding programs. This type of horizontal resistance is reported to be effective over space and time. Classical breeding techniques supported by marker-assisted selection (MAS) are the main tools in breeding programs. The aim of this study was to assess the possibility of transfer of slow-rusting genes from resistant genotypes of wheat into hexaploid triticale through cross-hybridizations. A total of 5,094 manual pollinations were conducted between two triticale cultivars Fredro and Twingo and 33 accessions of common wheat, which were reported as sources of slow-rusting resistance genes. The investigation of the slow-rusting gene transmission was performed using both molecular markers analyses and genomic in situ hybridization (GISH). In total, 34 F1 hybrid plants were obtained, and 29 of them carried both slow-rusting loci. Therefore, these hybrids may be used for triticale prebreeding program.

CERNBox is the CERN cloud storage hub for more than 25,000 users at CERN. It allows synchronising and sharing files on all major desktop and mobile platforms (Linux, Windows, MacOSX, Android, iOS) providing universal, ubiquitous, online- and offline access to any data stored in the CERN EOS infrastructure. CERNBox also provides integration with other CERN services for big science: visualisation tools, interactive data analysis and real-time collaborative editing.Over the last two years, CERNBox has evolved from a pure cloud sync and share platform into a collaborative service, to support new applications such as DrawIO for diagrams and organigrams sketching, OnlyOffice and Collabora Online for documents editing, and DXHTML Gantt for project management, as alternatives to traditional desktop applications. Moving to open source applications has the advantage to reduce licensing costs and enables easier integration within the CERN infrastructure. This move from commercial software to open source solutions is part of the MALT project, led by the IT department at CERN to reduce the dependencies on commercial solutions.As part of the MALT project, CERNBox is the chosen solution to replace Home directories of the Windows DFS file system. Access to storage from Windows managed devices for end-users is largely covered by synchronization clients. However, online access using standard CIFS/SMB protocol is required for shared use-cases, such as central login services (Terminal Services) and visitor desktop computers. We present recent work to introduce a set of Samba gateways running in High Availability cluster mode to enable direct access to the CERNBox backend storage (EOS).

A pivotal-differential evolution pattern is when two allopolyploids share a common genome, which is called pivotal, and differ with respect to the other genome or genomes, called differential. This feature induces the intergenomic recombination between chromosomes of differential genomes, which can lead to speciation. Our study is a cytomolecular insight into this mechanism which was adapted for the induction of intergenomic chromosome recombination in hybrids of synthetic amphidiploids Aegilops biuncialis × S. cereale (UUMMRR) and triticale (AABBRR) where R-genome was pivotal. We observed chromosome recombination events which were induced by both: (1) random chromosome fragmentation and non-homologous chromosome end joining at mitosis of root meristem cells and (2) intergenomic chromosome associations at meiosis of pollen mother cells (PMCs) of F1 hybrids. Reciprocal chromosome translocations were identified in six F1 plants and 15 plants of F2 generation using fluorescence in situ hybridization (FISH) with DNA clones (pTa-86, pTa-k374, pTa-465, pTa-535, pTa-k566, and pTa-713). We observed signals of pTa-86, pTa-535, and pTa-k566 probes in several chromosome breakpoints. The comparison of the DNA clone sequences distinguished a number of common motifs, which can be considered as characteristics of chromosome breakpoint loci. Immunodetection of synaptonemal complex proteins and genomic in situ hybridization analysis at meiosis of PMCs of F1 hybrids showed, that the homologous pairing of pivotal R—genome chromosomes is crucial for the fertility of F1 hybrids, however, these chromosomes can be also involved in the intergeneric recombination.

Segregation distorters are curious, evolutionarily selfish genetic elements, which distort Mendelian segregation in their favor at the expense of others. Those agents include gametocidal factors (Gc), which ensure their preferential transmission by triggering damages in cells lacking them via chromosome break induction. Hence, we hypothesized that the gametocidal system can be adapted for chromosome manipulations between Triticum and Secale chromosomes in hexaploid triticale (×Triticosecale Wittmack). In this work we studied the little-known gametocidal action of a Gc factor located on Aegilops geniculata Roth chromosome 4Mg. Our results indicate that the initiation of the gametocidal action takes place at anaphase II of meiosis of pollen mother cells. Hence, we induced androgenesis at postmeiotic pollen divisions (via anther cultures) in monosomic 4Mg addition plants of hexaploid triticale (AABBRR) followed by production of doubled haploids, to maintain the chromosome aberrations caused by the gametocidal action. This approach enabled us to obtain a large number of plants with two copies of particular chromosome translocations, which were identified by the use of cytomolecular methods. We obtained 41 doubled haploid triticale lines and 17 of them carried chromosome aberrations that included plants with the following chromosome sets: 40T+Dt2RS+Dt2RL (5 lines), 40T+N2R (1), 38T+D4RS.4BL (3), 38T+D5BS-5BL.5RL (5), and 38T+D7RS.3AL (3). The results show that the application of the Gc mechanism in combination with production of doubled haploid lines provides a sufficiently large population of homozygous doubled haploid individuals with two identical copies of translocation chromosomes. In our opinion, this approach will be a valuable tool for the production of novel plant material, which could be used for gene tracking studies, genetic mapping, and finally to enhance the diversity of cereals.

Three sets of hexaploid introgressive triticale lines, with Triticum monococcum ssp. monococcum (cultivated einkorn wheat) genes and a bread wheat chromosome 1D substituted for chromosome 1A, and one set of secondary triticale lines were evaluated for grain and flour physicochemical and dough rheological characteristics in two generations (F7 and F8). Genomic in situ hybridization (GISH) and fluorescence in situ hybridization (FISH) confirmed the 1D/1A chromosome substitution. The presence or absence of einkorn high-molecular-weight (HMW) glutenin subunits and the wheat Glu-D1d locus encoding the 5 + 10 subunits was assessed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), capillary zone electrophoresis, and allele-specific molecular markers. Significant differences were found among physicochemical properties (with the exception of the Hagberg falling number) of all introgressive Triticale/T. monococcum lines and the secondary triticale lines. The wheat 1D/1A chromosome substitution also affected these properties. The results showed that in all introgressive triticale lines, the protein and gluten content, Zeleny sedimentation value, and water absorption capacity, were increased. The rheological parameters estimated using micro-farinograph, reomixer, and Kieffer dough extensibility systems also showed an appreciable increase in dough-mixing properties, maximum resistance to extension (Rmax), and dough extensibility. Introgressive Triticale/T. monococcum lines with 5 + 10 subunits have particularly favorable rheological parameters. The results obtained in this study suggest that the cultivated einkorn genome Am, in the context of hexaploid secondary triticale lines and with a wheat 1D/1A substitution, has the potential to improve gluten polymer interactions and be a valuable genetic resource for triticale quality improvement.

Highlight: The level of pathogen alpha-amylase and plant beta-amylase activities could be components of plant-pathogen interaction associated with the resistance of triticale to Fusarium head blight. Triticale was used here as a model to recognize new components of molecular mechanism of resistance to Fusarium head blight (FHB) in cereals. Fusarium-damaged kernels (FDK) of two lines distinct in levels of resistance to FHB were applied into a proteome profiling using two-dimensional gel electrophoresis (2-DE) to create protein maps and mass spectrometry (MS) to identify the proteins differentially accumulated between the analyzed lines. This proteomic research was supported by a measurement of alpha- and beta-amylase activities, mycotoxin content, and fungal biomass in the analyzed kernels. The 2-DE analysis indicated a total of 23 spots with clear differences in a protein content between the more resistant and more susceptible triticale lines after infection with Fusarium culmorum. A majority of the proteins were involved in a cell carbohydrate metabolism, stressing the importance of this protein group in a plant response to Fusarium infection. The increased accumulation levels of different isoforms of plant beta-amylase were observed for a more susceptible triticale line after inoculation but these were not supported by a total level of beta-amylase activity, showing the highest value in the control conditions. The more resistant line was characterized by a higher abundance of alpha-amylase inhibitor CM2 subunit and simultaneously a lower activity of alpha-amylase after inoculation. We suggest that the level of pathogen alpha-amylase and plant beta-amylase activities could be components of plant-pathogen interaction associated with the resistance of triticale to FHB.

Triticum genus encloses several tetraploid species that are used as genetic stocks for expanding the genetic variability of wheat (Triticum aestivum L.). Although the T. aestivum (2n = 6x = 42, AABBDD) and T. durum (2n = 4x = 28, AABB) karyotypes were well examined by chromosome staining, Giemsa C-banding and FISH markers, other tetraploids are still poorly characterized. Here, we established and compared the fluorescence in situ hybridization (FISH) patterns on chromosomes of 20 accessions of T. polonicum species using different repetitive sequences from BAC library of wheat 'Chinese Spring'. The chromosome patterns of Polish wheat were compared to tetraploid (2n = 4x = 28, AABB) Triticum species: T. durum, T. diccocon and T. turanicum, as well. A combination of pTa-86, pTa-535 and pTa-713 probes was the most informative among 6 DNA probes tested. Probe pTa-k374, which is similar to 28S rDNA sequence enabled to distinguish signal size and location differences, as well as rDNA loci elimination. Furthermore, pTa-465 and pTa-k566 probes are helpful for the detection of similar organized chromosomes. The polymorphisms of signals distribution were observed in 2A, 2B, 3B, 5B, 6A and 7B chromosomes. Telomeric region of the short arm of 6B chromosome was the most polymorphic. Our work is novel and contributes to the understanding of T. polonicum genome organization which is essential to develop successful advanced breeding strategies for wheat. Collection and characterization of this germplasm can contribute to the wheat biodiversity safeguard.

The main aim of this work was to induce the chromosome rearrangements between Aegilops ovata (UUMM) and hexaploid triticale (AABBRR) by expression of the gametocidal factor located on the chromosome 4M. The Aegilops ovata × Secale cereale (UUMMRR) amphiploids and triticale ‘Moreno’ were used to produce hybrids by reciprocal crosses. Chromosome dynamics was observed in subsequent generations of hybrids during mitotic metaphase of root meristems and first metaphase of meiosis of pollen mother cells. Chromosomes were identified by genomic in situ hybridisation (GISH) and fluorescence in situ hybridisation (FISH) using pTa71, pTa791, pSc119.2 and pAs1 DNA probes. It has been shown that the origin of the genetic background had an influence on Aegilops chromosome transmission. Moreover, it has been reported that the preferential transmission of chromosome 4M appeared during both androgenesis and gynogenesis. It is also hypothesised that the expression of the triticale Gc gene suppressor had an influence on the semi-fertility of hybrids but did not inhibit the chromosome rearrangements. This paper also describes the double haploid production, which enabled to obtain plants with two identical copies of triticale chromosomes with translocations of Aegilops chromatin segments.

Allocation of the chromosome 2D of Ae. tauschii in triticale background resulted in changes of its organization, what is related to varied expression of genes determining agronomically important traits. Monosomic alien addition lines (MAALs) are crucial for transfer of genes from wild relatives into cultivated varieties. This kind of genetic stocks is used for physical mapping of specific chromosomes and analyzing alien genes expression. The main aim of our study is to improve hexaploid triticale by transferring D-genome chromatin from Aegilops tauschii × Secale cereale (2n = 4x = 28, DDRR). In this paper, we demonstrate the molecular cytogenetics analysis and SSR markers screening combined with phenotype analysis and evaluation of powdery mildew infection of triticale monosomic addition lines carrying chromosome 2D of Ae. tauschii. We confirmed the inheritance of chromosome 2D from the BC2F4 to the BC2F6 generation of triticale hybrids. Moreover, we unveiled a high variable region on the short arm of chromosome 2D, where chromosome rearrangements were mapped. These events had direct influence on plant height of hybrids what might be connected with changes at Rht8 loci. We obtained 20 semi-dwarf plants of BC2F6 generation carrying 2D chromosome with the powdery mildew resistance, without changes in spike morphology, which can be used in the triticale breeding programs.