Isolation and evaluation of different wheat-rye translocation lines obtained from a disease resistant double translocation line with 1BL/1RS and 2RL/2BS

Wheat-rye translocations involving 1RS and 2RL of rye are the most useful sources of genes for disease resistance in wheat breeding. Rye genes are known to control resistance to biotic and abiotic stresses. Wheat-rye translocations have been widely used by breeders all over the world because genes located on translocated chromosome arms or fragments from the rye genome can determine a number of useful traits in wheat, such as high yield, wide adaptation, diseases and pest resistance. The wheat-rye translocation lines used in this study were derived from a cross between the Swedish bread wheat variety Topper and the line KR99-139 being homozygous for the two different wheat-rye translocations 1BL/1RS and 2RL/2BS. BC1F1 materials were obtained through one back-cross with either the line KR99-139 or the variety Topper. Thereafter, BC1F2 and BC1F3 were obtained by once and twice selfing. In the obtained material, it was thereafter possible to define four different possible homozygous translocation combinations. Thus, lines containing both 1RS and 2RL translocations, containing only 1RS or 2RL and without any translocation were identified. For identification of the four possible homozygous wheat-rye translocation lines mentioned above, three different methods were used. First, lines of different types were characterized and isolated based on a phenotypical marker, i.e. if the plant showed red or green coleoptile colour. Plants with homozygous presence of 2RL were known to develop red coleoptile, as a gene for red coleoptile has been verified to be present at 2RL in these lines. The analyses of coleoptile colours were done in the BC1F2 (obtained from selfed BC1F1 lines determined by molecular markers at BAZ, Germany to be 1RS– –/2RL+–) and BC1F3 (obtained from the BC1F2 lines having a red coleoptile) wheat-rye translocation lines, where the variety Topper had been used for backcrossing. Moreover, the BC1F2 (obtained from selfed BC1F1 lines determined by molecular markers at BAZ, Germany to be 1RS++/2RL+–) wheat-rye translocation lines for which the KR99-139 line was used for backcrossing, was selfed, and analyses of coleoptile colours were done in the BC1F3 (on a representative sample of all combinations of presence and absence of 2RL). The results from the coleoptiles colour analyses generally showed that it was possible to distinguish lines having 2RL++ (red coleoptiles) and 2RL– – (green coleoptiles). 7 Plants having 2RL+– were sometimes classified as having green and sometimes as having red coleoptiles. Therefore, if coleoptiles colour is going to be used for selection of lines with presence/absence of 2RL in homozygous form, at least two generations have to be analyzed and lines not segregating in either of the analyses can be judged as being homozygous as related to their coleoptiles colour. For identification of lines with presence of heterozygous 1RS+– and homozygous 2RL++ rye chromosome the Giemsa C-banding technique was used. The Giemsa C-banding techniques on the BC1F3 segregating population generally resulted in well-defined sharp, distinct bands in the wheat-rye translocation lines and both the rye chromosome arms, 1RS and 2RL were identified. Additionally, five microsatellite (SSR) markers SCM9, SCM39, SCM43, SCM69 and SCM75 were used for verification of the presence of 1RS and 2RL. Among the five SSR markers, SCM9 and SCM75 resulted in reliable amplification of expected products, 220 bp and 191 bp respectively. The line KR99-139 containing both 1RS and 2RL showed correct amplification products with both mentioned primers while the bread wheat variety Topper without any rye chromosome showed no amplification with both SSR primers pairs. Resistance towards yellow rust and stem rust were evaluated through seedling resistance test in the greenhouse (Global Rust Reference Center, Denmark) to Puccinia striiformis, and adult plant resistance to Puccinia graminis, race Ug99 (TTKSK) in Njoro, Kenya. For the seedling resistance test, pathogenicity of 17 races/isolates of yellow rust was used. The BC1F3 which carries combination of 1RS++/2RL++, 1RS++/2RL+– and the KR99-139 were found to be highly resistant to some races/isolates whether the variety Topper was fully susceptible to all races/isolates. The results showed Yr9 to be one possible gene that could be responsible for the obtained yellow rust resistance. However, the results were not that clear so than not other possible genes could also be an alternative. For adult plant resistance towards Ug99, a total of 28 of the BC1F3 wheat-rye translocation lines and their parents were evaluated in the field of Njoro, Kenya. The results indicated that out of the 30 tested lines 20 were susceptible, 8 moderately susceptible to susceptible and in 2 lines the resistance to Ug99 was identified. The two BC1F3 wheat-rye translocation lines that were found to be resistant towards Ug99 were both being homozygous for 1RS++ and heterozygous for 2RL+–. Thus, these results indicated presence of several genes/QTLs controlling resistance indicating possible epistatic effects of the genes involved. The lines identified as resistant will be utilized in combination with Tajik germplasm 8 to develop a mapping population for determining the underlying basis of resistance. To summarize results from the research outlined in this thesis indicate that wheat-rye translocation lines and used methods can be highly relevant for wheat breeding programs and further research

Genetic characterisation of novel resistance alleles to stem rust and stripe rust in wheat-alien introgression lines

Bread wheat (Triticum aestivum L., 2n = 6x = 42, AABBDD) is one of the most important food crops world-wide, but is attacked by many diseases and pests that cause significant yield losses. Globally, stem rust (Sr) (Puccinia graminis f. sp. tritici Erikss & E. Henning), stripe rust (Yr) (Puccinia striiformis Westend. f. sp. tritici Eriks) and leaf rust (Lr) (Puccinia triticina Eriks) are a great threat to wheat production. The majority of the Sr, Yr and Lr resistance genes are already defeated by numerous virulent races, so enhanced genetic resistance against these devastating diseases are essential. Wheat-alien introgressions from derivatives of Secale cereale L. (2n = 2x = 14, RR), Leymus mollis (2n = 4x = 28, NsNsXmXm), Leymus racemosus (2n = 4x = 28, NsXm) and Thinopyrum junceiforme (2n = 4x = 28; J1J1J2J2) are important genetic resources for new sources of resistance genes. To identify new sources of resistance, this thesis evaluated seedling and adult plant resistance to a wide array of stem rust and stripe rust races. Three wheat-rye disomic substitution lines 2R (2D) were found to carry new resistance gene/s to stem rust races and six multiple wheat-rye introgression lines with 5RS·5AL+4R+6R carried new resistance gene/s to stripe rust races. At adult plant stage, the wheat-rye translocation line with 1BL·1RS and 2RL·2BS exhibited low susceptibility to race TTKSK under field conditions. The wheat-rye T2DS·2RL Robertsonian translocation line (TA5094) with a new stem rust resistance gene was developed through the breakage-fusion mechanism and verified using seedling resistance assays and molecular and cytogenetic analyses. Three kompetitive allele-specific PCR (KASP) markers located on rye chromosome 2RL were identified as being closely associated with the new stem rust resistance gene. Fluorescence in situ hybridisation (FISH) analysis confirmed the resistance gene in F3:4 homozygous lines. The stem rust resistance gene in TA5094 line on chromosome 2RL arm was designated Sr59. Wheat cultivars, advanced lines and landraces from Tajikistan were assessed at seedling and adult plant stages against Sr, Yr and Lr races. Based on multipathotype assessment and molecular markers, the presence of Sr6, Sr31/Yr9/Lr26, Sr38/Yr17/Lr37, Yr2 and Yr27 and pleiotropic resistance genes Sr57/Lr34/Yr18/ and Sr2/Yr30/Lr27 was postulated. Overall, this thesis identified novel genetic resistance resources against stem rust, stripe rust and leaf rust in Tajik wheat and in wheat-alien introgressions. This resistance gene/s will be useful in diversifying the current set of resistance genes deployed to control these devastating diseases

Developing adapted wheat lines with broad-spectrum resistance to stem rust: Introgression of Sr59 through backcrossing and selections based on genotyping-by-sequencing data

Control of stem rust, caused by Puccinia graminis f.sp. tritici, a highly destructive fungal disease of wheat, faces continuous challenges from emergence of new virulent races across wheat-growing continents. Using combinations of broad-spectrum resistance genes could impart durable stem rust resistance. This study attempted transfer of Sr59 resistance gene from line TA5094 (developed through CSph1bM-induced T2DS center dot 2RL Robertsonian translocation conferring broad-spectrum resistance). Poor agronomic performance of line TA5094 necessitates Sr59 transfer to adapted genetic backgrounds and utility evaluations for wheat improvement. Based on combined stem rust seedling and molecular analyses, 2070 BC1F1 and 1230 BC2F1 plants were derived from backcrossing BAJ#1, KACHU#1, and REEDLING#1 with TA5094. Genotyping-by-sequencing (GBS) results revealed the physical positions of 15,116 SNPs on chromosome 2R. The adapted genotypes used for backcrossing were found not to possess broad-spectrum resistance to selected stem rust races, whereas Sr59-containing line TA5094 showed resistance to all races tested. Stem rust seedling assays combined with kompetitive allele-specific PCR (KASP) marker analysis successfully selected and generated the BC2F2 population, which contained the Sr59 gene, as confirmed by GBS. Early-generation data from backcrossing suggested deviations from the 3:1 segregation, suggesting that multiple genes may contribute to Sr59 resistance reactions. Using GBS marker data (40,584 SNPs in wheat chromosomes) to transfer the recurrent parent background to later-generation populations resulted in average genome recovery of 71.2% in BAJ#1*2/TA5094, 69.8% in KACHU#1*2/TA5094, and 70.5% in REEDLING#1*2/TA5094 populations. GBS data verified stable Sr59 introgression in BC2F2 populations, as evidenced by presence of the Ph1 locus and absence of the 50,936,209 bp deletion in CSph1bM. Combining phenotypic selections, stem rust seedling assays, KASP markers, and GBS data substantially accelerated transfer of broad-spectrum resistance into adapted genotypes. Thus, this study demonstrated that the Sr59 resistance gene can be introduced into elite genetic backgrounds to mitigate stem rust-related yield losses

Genome-Wide Association Study Identifies Two Loci for Stripe Rust Resistance in a Durum Wheat Panel from Iran

Stripe rust (Puccinia striiformis f. sp. tritici (Pst)) is one of the most devastating fungal diseases of durum wheat (Triticum turgidum L. var. durum Desf.). Races of Pst with new virulence combinations are emerging more regularly on wheat-growing continents, which challenges wheat breeding for resistance. This study aimed to identify and characterize resistance to Pst races based on a genome-wide association study. GWAS is an approach to analyze the associations between a genome-wide set of single-nucleotide polymorphisms (SNPs) and target phenotypic traits. A total of 139 durum wheat accessions from Iran were evaluated at the seedling stage against isolates Pstv-37 and Pstv-40 of Pst and then genotyped using a 15K SNP chip. In total, 230 significant associations were identified across 14 chromosomes, of which 30 were associated with resistance to both isolates. Furthermore, 17 durum wheat landraces showed an immune response against both Pst isolates. The SNP markers and resistant accessions identified in this study may be useful in programs breeding durum wheat for stripe rust resistance

Stem Rust Resistance in 1BL.1RS and 2RL.2BS Double Wheat-Rye Translocation Lines

The wheat stem rust pathogen, Puccinia graminis f.sp. tritici, is a significant and devastating disease of wheat crops worldwide. Wheat has many wild relatives in which to source new resistance genes, including the cereal crop of rye in the tertiary genepool. The aim of this study was to assess the reaction of 1BL.1RS and 2RL.2BS double wheat-rye translocation lines to virulent stem rust races from Africa and North America. BC1F3 and BC1F4 populations from a cross between the line KR99-139 (a double wheat-rye translocation line with 1BL.1RS and 2RL.2BS) and the bread wheat cultivar Topper were used in the study. Several of the populations homozygous for 1BL.1RS and heterozygous for 2RL.2BS showed resistance and low severity adult plant resistance (20RMR-50MSS) to the African stem rust race TTKSK in the field. None of the tested populations with varying chromosome combinations showed seedling resistance to any of the tested stem rust races. Thus, these resistant populations likely carry gene/s effective at the adult plant stage since all stage resistance genes with major effect appear to be absent based on the seedling assays. Resistant lines combined three chromosomes (1RS, 2RS and 2BS) which make their direct use in breeding more complicated. Mapping studies followed by potential transfer of genes between 2R and 2B will make the identified minor genes more useful in wheat breeding

Sources of resistance to yellow rust and stem rust in wheat-alien introgressions

Wheat is the staple food and the main source of caloric intake in most developing countries, and thereby an important source in order to maintain food security for the growing populations in those countries. Stem rust Puccinia graminis f. sp. tritici, and yellow rust P. striiformis f. sp. tritici of wheat continues to cause severe damage locally and globally, thereby contributing to food insecurity. In this paper biology and taxonomy of stem rust and yellow rust, breeding for resistance, utilization of resistance sources from different gene pools, molecular characterization and genetic dissection of resistance to rusts are discussed

Diverse Wheat-Alien Introgression Lines as a Basis for Durable Resistance and Quality Characteristics in Bread Wheat

Wheat productivity has been significantly improved worldwide through the incorporation of novel genes from various gene pools, not least from wild relatives of wheat, into the commonly cultivated bread and durum wheat. Here, we present and summarize results obtained from a diverse set of wheat-alien introgression lines with mainly introgressions of rye, but also ofLeymusspp. andThinopyrum junceiformeinto bread-wheat (Triticum aestivumL.). From this material, lines carrying 2RL were found with good agronomic performance and multiple resistance not least towards several races of powdery mildew. A novel resistance gene, one of few showing resistance towards all today identified stem rust races, designatedSr59, was also found originating from 2RL. Lines with multiple introgressions from 4R, 5R, and 6R were found resistant towards the majority of the stripe rust races known today. Due to lack of agricultural adaptation in these lines, transfer of useful genes into more adapted wheat material is a necessity, work which is also in progress through crosses with the CSph1bmutant, to be able to only transfer small chromosome segments that carry the target gene. Furthermore, resistance towards Russian wheat aphid was found in lines having a substitution of 1R (1D) and translocations of 3DL.3RS and 5AL.5RS. The rye chromosomes 1R, 2R, and 6R were found responsible for resistance towards the Syrian Hessian fly. High levels of especially zinc was found in several lines obtained from crosses withLeymus racemosusandLeymus mollis, while also some lines with 1R, 2R, or 5R showed increased levels of minerals and in particular of iron and zinc. Moreover, lines with 1R, 2R, 3R, andLeymusspp. introgressions were also found to have a combination of high iron and zinc and low cadmium concentrations. High variation was found both in grain protein concentration and gluten strength, measured as %UPP, within the lines, indicating large variation in bread-making quality. Thus, our study emphasizes the impact that wheat-alien introgression lines can contribute to current wheat lines and shows large opportunities both to improve production, resistance, and quality. To obtain such improvements, novel plant breeding tools, as discussed in this paper, opens unique opportunities, to transfer suitable genes into the modern and adapted wheat cultivars

Recurrent parent genome recovery in the BC₂F₂ generation using 40,584 genome-wide SNPs.

Recurrent parent genome recovery in the BC2F2 generation using 40,584 genome-wide SNPs.</p

PCA plot of rye chromosome 2R using 15,116 SNPs from GBS reads.

A) Resistant parental lines (SLU238, SLU239, and TA5094) with chromosome 2R; B) BC2F2 population comprising recurrent parents carrying chromosome 2RL close to SLU238 and TA5094; C) BC2F2 population derived from recurrent parents carrying the chromosome 2RL segment; D) recurrent parents (BAJ #1, KACHU #1, and REEDLING #1) and the susceptible BC2F2 population without chromosome 2RL; E) lines CSph1bM and CSA.</p

Marker-assisted selection for the improvement of cereals and pseudocereals

Cereals are grasses (a monocot family Poaceae, also known as Gramineae) cultivated widely for their grains used in human and animal consumption. From ancient times, cereals have played an important role in world agriculture and nowadays their significance is illustrated by the overall production of 2.996 million tons being harvested globally in 2020 (FAOSTAT, 2021). The most important staple cereal crops are wheat, rice, maize, sorghum, barley, oats, and millet. However, wheat, rice, and maize together comprise at least 75% of the world’s grain production, with 761, 757, and 1.162 million tons harvested in 2020, respectively. Rice, sorghum, millet, and wheat are widely produced in Asia; corn and sorghum in America; and barley, rye, and oats in Europe. These three continents produce together 80% of the world’s cereal grains. Cereals are a pivotal nutrient source in both developed and developing countries since their grains contain major nutritional and energy sources (proteins, carbohydrates, minerals, amino acids, fibers) as well as micronutrients (vitamins, magnesium, and zinc) (O’Neil et al., 2010; Papanikolaou & Fulgoni, 2017). However, the utilization pattern of these cereal grains differs. In developed countries, more than 70% of total cereal production is fed to the animals, whereas in underdeveloped countries, 68%–98% of the cereal production is used for human consumption