Reference genome-assisted identification of stem rust resistance gene Sr62 encoding a tandem kinase

The wild relatives and progenitors of wheat have been widely used as sources of disease resistance (R) genes. Molecular identification and characterization of these R genes facilitates their manipulation and tracking in breeding programmes. We developed a reference-quality genome assembly of the wild diploid wheat relative Aegilops sharonensis and used positional mapping, mutagenesis, RNA-Seq and transgenesis to identify the stem rust resistance gene Sr62, which was also transferred to common wheat. This gene encodes a tandem kinase, homologues of which exist across multiple taxa in the plant kingdom. Stable Sr62 transgenic wheat lines showed high levels of resistance against diverse isolates of the stem rust pathogen, highlighting the utility of Sr62 for deployment as part of a polygenic stack to maximize the durability of stem rust resistance.

DIVERSITY OF WIDE HYBRIDIZATION DERIVATIVES OF WINTER WHEAT FOR RESISTANCE TO DISEASES AND OTHER ALIEN CHARACTERS

Introduction. The global climate change conduces spreading of main diseases of winter bread wheat (Triticum aestivum L.) and increases the yield losses caused by both these diseases and drought. Preventing the devastating impact of these factors on the yield depends on development of donors carrying the resistance genes. Aim. The diversity of new introgression wheat lines derived from complex interspecies crosses was evaluated for resistance to widespread diseases, drought and other agricultural traits. Methods. The trial was conducted consecutively during two vegetative periods (2018-2019 and 2019-2020) on the black earth area under the arid system in a control nursery design. Genetic material includes seventy-eight introgression wheat lines of different origin with two check cultivars for the arid climate zone of Ukraine. The data on nine agronomic (quantitative scores) and five plant pathological (point scores) traits were collected and subjected to both variance and correlation analysis to comprehend the contribution of the factors towards general diversity and connections between the characters. Results. Most lines were resistant to rust species due to the successful introgression of alien Lr, Yr and Sr genes from all sources involved in hybridization. High long-term resistance to stem rust was observed mainly among the derivatives of the collection sample H74/90-245 from Bulgaria, whose genetic background was favorable for generating a positive effect of 1BL.1RS translocation on both productive and adaptive traits in southern Ukraine and the ability to combine with other resistance genes. The yield traits did not correlate with the resistance to diseases except stem rust (Rsp = 0.34**) and Septoria blight (Rsp = −0.23*). Resistance to stem rust correlated with the grain and protein yield, test weight and flour density, which may be due to the presence of translocation 1BL.1RS. Negative correlation was observed between quality traits and grain yield, but quality traits mainly positively correlated with each other. The plant height had the highest contribution to grain yield followed by volume of 1000 kernels and WTK. Conclusions. The introgression lines were found to be diverse and potential for use in the wheat improvement programs for resistance to the diseases or drought in Southern Ukraine.

Virulence Structure of the Wheat Powdery Mildew Population in Serbia

Powdery mildew is a common, economically important disease in the wheat growing area of Serbia. A large-scale virulence survey of its causal agent Blumeria graminis f. sp. tritici population was performed in the period 1995–2013. A total of 1013 isolates were recovered from the collected chasmothecial samples. Among them, 862 unique pathotypes were identified using a differential set of 20 wheat lines with known powdery mildew (Pm) resistant genes. The pathogen was highly diverse. Number of virulence genes (virulence complexity) per isolate was large, supporting a constant need to extend the differential set of wheat with newly identified Pm genes. Virulence frequencies to Pm6, Pm7, and Pm5+8 were high throughout the 19-year period, in contrast with that to Pm5+6, which was consistently at a low level. The most significant change in the population was observed for virulence to the Pm2+4b+6 gene combination, with an increasing frequency of virulence to this gene combination over the years. High virulence complexity and genetic diversity of the population are the most influential factors for the damaging epidemics that this pathogen can cause.

Molecular labeling of Vrn, Ppd genes and vernalization response of the ultra-early lines of spring bread wheat Triticum aestivum L.

Background. The knowledge of genetic control of vernalization response in the ultra-early accessions can facilitate bread wheat breeding for a high adaptive capacity. Materials and methods. The study involved the ultra-early lines Rico (k-65588) and Rimax (k-67257) as the earliest maturing lines in the VIR bread wheat collection, as well as 10 Rifor lines (k-67120, k-67121, k-67250-67256) with a high rate of development before heading. A late ripening accession ‘Forlani Roberto’ (k-42641) and ‘Leningradskaya 6’ variety (k-64900), regionally adapted to Northwestern Russia, were also studied. The alleles of the Vrn and Ppd genes were identified by the PCR analysis using the allele-specific primers published in literature sources. The response to vernalization (30 days at 3°C) and a short 12-hour day were determined using a methodology accepted at VIR. Results. The ultra-early lines respond to a short 12-hour day and 30-day vernalization very poorly. The genotype of ultra-early wheat lines is mainly represented by three genes, Vrn-A1, Vrn-B1a, and Vrn-D1, which ensure insensitivity to vernalization alongside with the expression of Ppd-D1a, which controls the response to photoperiod. The ultra-early lines Rifor 4 and Rifor 5 have a recessive allele vrn-A1a, like the original ‘Forlani Roberto’ accession. The lines Rifor 4 and Rifor 5 are vernalization-insensitive under the long day and have a very weak response under the short day (3.5±0.42 days and 4.0±0.61 days, respectively). However, ‘Forlani Roberto’ with the vrn-A1a gene responds to vernalization in the same way under any photoperiod (12.3±1.58 days and 12.2±0.74 days). Conclusion The ultra-early lines of bread wheat Rifor 4 and Rifor 5 with the vrn-A1a gene can have no response to vernalization or have a low level response. This effect can be a reason for the formation of a complex of modifier genes along with the dominant gene Vrn-D1, which forms during the hybridization of F7-8 Rico × Forlani Roberto. The ultra-early lines of bread wheat Rico, Rimax and Rifor (k-67120, k-67121, k-67250-67256) can serve as effective sources of genes for earliness in common wheat breeding.

Exploring the Genetic Variability for Yield Attributing Traits among the Indigenous and Exotic Collection of Wheat in Cis-Himalayan Region of West Bengal, India

A field experiment was conducted at Instructional Farm, Uttar Banga Krishi Viswavidyalaya, during the 2018-19 rabi season to study the genetic components and genetic variability based on eleven number of quantitative traits in 254 bread wheat lines, which were collected from NBPGR, New Delhi. Analysis of variance for treatment showed high significance for all the traits except grain yield polt-1. Estimated GCV and PCV were high for awn length, peduncle length, plant height and AUDPC. Plant height (97.83), awn length (95.49), peduncle length (94.88), test weight (90.98), days to 50% flowering (87.66), number of grains spike-1 (70.33) and spike length (62.3) showed high heritability along with a high degree of genetic advance. Finally, 254 wheat accessions were grouped into three clusters based on the dendrogram analysis using the ward method.

A prebreeding study of introgression spring bread wheat lines carrying combinations of stem rust resistance genes, Sr22+Sr25 and Sr35+Sr25

The Sr22, Sr35, and Sr25 genes attract the attention of bread wheat breeders with their effectiveness against Puccinia graminis f. sp. tritici race Ug99 and its biotypes. The effectiveness and impact of Sr22+Sr25 and Sr35+Sr25 gene combinations on agronomic traits have not yet been studied. In the present article, these traits were studied using the spring bread wheat lines L503/W3534//L503, L503/Sr35//L503/3/L503 carrying the Sr22+Sr25 and Sr35+Sr25 genes during 2016–2020. These lines were assessed for resistance to P. graminis f. sp. tritici under natural epiphytotics and to the Saratov, Lysogorsk and Omsk populations of the pathogen and to the PgtZ1 (TKSTF) and PgtF18.6 fungus isolates in laboratory conditions (TKSTF + Sr33). The presence of the studied Sr-genes was confirmed by using molecular markers. Prebreeding studies were conducted during 2018–2020 vegetation periods. Under the natural epiphytotics of the pathogen and in the laboratory conditions, the Sr22+Sr25 combination was highly effective, while Sr35+Sr25 was ineffective. For grain yield, the lines with the Sr22+Sr25 and Sr35+Sr25 genes were superior to the recipient cultivar L503 in one year (Sr22+Sr25 in 2019; Sr35+Sr25 in 2018), with a decrease in 2020, but in general there were no differences. For the period 2018–2020, both combinations showed a decrease in 1000 grains weight and an increase in the germination-earing period. The line with Sr22+Sr25 genes showed insignificant effects on gluten and dough tenacity, but the ratio of dough tenacity to extensibility was higher, and flour strength, porosity and bread volume were lower; in the line with Sr35+Sr25 genes, the gluten content was lower, but the strength, tenacity of the dough and the ratio of dough tenacity to extensibility were higher, flour strength and the porosity of the bread were at the recipient level, but the volume of bread was lower.

A Bioinformatic Workflow for InDel Analysis in the Wheat Multi-Copy α-Gliadin Gene Family Engineered with CRISPR/Cas9

The α-gliadins of wheat, along with other gluten components, are responsible for bread viscoelastic properties. However, they are also related to human pathologies as celiac disease or non-celiac wheat sensitivity. CRISPR/Cas was successfully used to knockout α-gliadin genes in bread and durum wheat, therefore, obtaining low gluten wheat lines. Nevertheless, the mutation analysis of these genes is complex as they present multiple and high homology copies arranged in tandem in A, B, and D subgenomes. In this work, we present a bioinformatic pipeline based on NGS amplicon sequencing for the analysis of insertions and deletions (InDels) in α-gliadin genes targeted with two single guides RNA (sgRNA). This approach allows the identification of mutated amplicons and the analysis of InDels through comparison to the most similar wild type parental sequence. TMM normalization was performed for inter-sample comparisons; being able to study the abundance of each InDel throughout generations and observe the effects of the segregation of Cas9 coding sequence in different lines. The usefulness of the workflow is relevant to identify possible genomic rearrangements such as large deletions due to Cas9 cleavage activity. This pipeline enables a fast characterization of mutations in multiple samples for a multi-copy gene family.

Response of durum wheat lines to the cause agent of leaf rust Puccinia triticina

Аbstract. The response of 1262 durum wheat lines to the cause agent of leaf rust Puccinia triticina was studied. The investigation was carried under infection field conditions during 2013-2015 at Dobrudzha Agricultural Institute (DAI) – General Toshevo, Bulgaria. Over the years of investigation, a maximum high infection background was developed by the cause agent of the disease, including all pathogen pathotypes identified during this period. The race variability in the population of the pathogen during the period was determined at the Plant Pathology Laboratory of DAI according to a validated methodology for working with rusts. Seventy-one phenotypically different virulent pathotypes were established. Fifteen new races of P. triticina were also identified, which have not been present during the previous 15 years in Bulgaria. Seventeen lines and one cultivar with a certain degree of resistance to the pathogen were selected. The selected lines can be involved in the breeding programs for developing initial material with resistance to leaf rust P. triticina.

Assessing the Rheological Properties of Durum Wheat Semolina: A Review

Empiric rheology is considered a useful tool for assessing the technological quality of wheat. Over the decades, several tests have been adapted from common to durum wheat, and new approaches have been proposed to meet the needs of the players of the durum wheat value chain. Breeders are looking for reliable methods to test the functional quality of wheat lines at early stages, where there are limited amounts of sample; millers need fast and reliable methods for checking wheat quality right at the point of the receiving station; and pasta-makers are looking for suitable methods to predict end product quality. This review provides an overview of the strengths and weaknesses of the rheological tests currently used to evaluate the quality of durum wheat semolina, with the emphasis on Europe. Moreover, the relationships among the parameters obtained from different rheological approaches are extrapolated from the literature and integrated with the data obtained from 74 samples of durum wheat semolina. Although numerous efforts have been made to propose rapid and reliable tests for semolina characterization, the ideal test has yet to be proposed, indicating that researchers and pasta companies need to focus on perfecting the way to assess the quality of durum wheat and pasta.

Wheat Apoplast-Localized Lipid Transfer Protein TaLTP3 Enhances Defense Responses Against Puccinia triticina

Plant apoplast serves as the frontier battlefield of plant defense in response to different types of pathogens. Many pathogenesis-related (PR) proteins are accumulated in apoplastic space during the onset of plant–pathogen interaction, where they act to suppress pathogen infection. In this study, we found the expression of Triticum aestivum lipid transfer protein 3 (TaLTP3) gene was unregulated during incompatible interaction mediated by leaf rust resistance genes Lr39/41 at the early infection stage. Stable transgenic wheat lines overexpressing TaLTP3 exhibited enhanced resistance to leaf rust pathogen Puccinia triticina. Transcriptome analysis revealed that overexpression of TaLTP3 specifically activated the transcription of pathogenesis-related protein 1a (TaPR1a) and multiple plant hormone pathways, including salicylic acid (SA), jasmonic acid (JA), and auxin, in response to the infection of the model bacterial pathogen Pseudomonas syringae pv. tomato DC3000. Further investigation indicated that TaLTP3 physically associated with wheat TaPR1a protein in the apoplast. Transgenic wheat lines overexpressing TaLTP3 and TaPR1a showed higher accumulations of reactive oxygen species (ROS) during plant defense responses. All these findings suggested that TaLTP3 is involved in wheat resistance against leaf rust pathogen infection and forming a TaLTP3-TaPR1a complex in apoplast against this pathogen, which provides new insights into the functional roles of PR proteins.