Dr. Brian Steffenson

Persistent link for this collectionhttps://hdl.handle.net/11299/184255

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Aegilops sharonensis: Origin, genetics, diversity, and potential for wheat improvement
(Botany, 2009-08) Steffenson, Brian; Olivera, Pablo D.
Aegilops sharonensis Eig (Sharon goatgrass; section Sitopsis) is an annual diploid grass species growing endemically in the coastal plains of Israel and southern Lebanon. It is a wild relative of wheat, with a genome closely related to the B genome of cultivated bread wheat. With the most limited distribution of any species in the genus Aegilops, Ae. sharonensis is rapidly losing its habitats, owing to the combined effects of modern agricultural intensification and expansion of urban and industrial areas. Aegilops sharonensis is known to be a rich source of genes providing resistance to important wheat diseases and abiotic stresses, but it has not been widely exploited. The presence of gametocidal genes that control preferential transmission of chromosome 4Ssh increases the difficulty of introgressing genes from Ae. sharonensis into wheat. However, successful introgression of the genes for resistance to leaf rust, stripe rust, and powdery mildew has been achieved. Studies on genetic and phenotypic diversity indicated that Ae. sharonensis is a highly diverse species, comparable with others that have a wider geographic distribution and more variable environments. Targeting the regions and sites with the highest diversity in Ae. sharonensis will facilitate the capture of the greatest variability and also the identification of novel and diverse genes for wheat improvement.
Agronomic characteristics, malt quality, and disease resistance of barley germplasm lines with partial Fusarium head blight resistance
(Crop Science, 2005-07) Steffenson, Brian; Urrea, Carlos A.; Horsley, Richard D.; Schwarz, Paul B.
Fusarium head blight (FHB), incited by Fusarium graminearum Schwabe, has caused devastating losses in both yield and quality of barley (Hordeum vulgare L.) produced in the northern Great Plains from 1993 to 2003. Thirty-five barley germplasm lines with partial resistance to FHB have been identified in exotic and unadapted germplasm lines. Little is known about their agronomic characteristics, malt quality, and reaction to other diseases as compared to adapted cultivars. This information is needed so barley breeders can make informed decisions when planning crosses involving the resistant germplasm lines. The objective of this study was to compare the agronomic performance, malt quality, and disease reaction of barley germplasm lines with partial FHB resistance to cultivars grown in the northern Great Plains. Agronomic and malting data were collected on the 35 germplasm lines and five check cultivars grown in five environments in North Dakota from 1998 to 2000. Data for FHB severity and deoxynivalenol (DON, a mycotoxin produced by F. graminearum) accumulation were obtained for the same 40 entries grown in FHB-epidemic nurseries in North Dakota from 1997 to 1999. Seedling responses to foliar pathogens common in the northern Great Plains were determined in the greenhouse during fall 1997. None of the FHB-resistant barley germplasm lines had acceptable malt quality for all traits. Kernel plumpness, grain protein concentration, and malt extract were the traits impacted most severely. The FHB-resistant barley germplasm lines headed significantly later than the adapted barley cultivars. Most FHB-resistant germplasm lines were susceptible to the common foliar diseases of the northern Great Plains. At least four cycles of breeding will probably be necessary to develop FHB-resistant germplasm lines acceptable to producers and the malting and brewing industry.
Amplified fragment length polymorphism and virulence polymorphism in Puccinia hordei
(Canadian Journal of Plant Pathology, 2007) Sun, Y; Neate, S.M.; Zhong, S; Steffenson, Brian; Friesen, T. L.
Puccinia hordei is the causal agent of barley leaf rust. To study the genetic diversity in P. hordei, 45 isolates with diverse virulence patterns and geographical origins were analyzed using amplified fragment length polymorphism markers. Two pathotypes of Puccinia graminis f. sp. tritici and one isolate of P. graminis f. sp. secalis were included in the analysis for comparison. Six primer-pair combinations of amplified fragment length polymorphism were used and a total of 782 polymorphic markers were generated. Cluster analysis showed that P. graminis f. sp. tritici and P. graminis f. sp. secalis were distinctly different from P. hordei. The P. hordei isolates were clustered into five groups: group I contained a single, rare isolate that was virulent on all resistance genes except Rph13 and Rph15 ; group II contained a single isolate found to be virulent on the resistance gene Rph15 ; group III contained 2 isolates; group IV contained 24 isolates, 11 from the United States and 13 from diverse locations around the world; and group V contained 17 isolates, 7 from California, 7 from other states of United States, and 3 from central Europe. The study revealed that molecular diversity in P. hordei can be associated with virulence, but not well with geographic origin.
Analysis of ergosterol in single kernel and ground grain by gas chromatography-mass spectrometry
(Journal of Agricultural and Food Chemistry, 2006-05-11) Steffenson, Brian; Dong, Yanhong; Mirocha, Chester J
A method for analyzing ergosterol in a single kernel and ground barley and wheat was developed using gas chromatography−mass spectrometry (GC-MS). Samples were saponified in methanolic KOH. Ergosterol was extracted by “one step” hexane extraction and subsequently silylated by N-trimethylsilylimidazole/trimethylchlorosilane (TMSI/TMCS) reagent at room temperature. The recoveries of ergosterol from ground barley were 96.6, 97.1, 97.1, 88.5, and 90.3% at the levels of 0.2, 1, 5, 10, and 20 μg/g (ppm), respectively. The recoveries from a single kernel were between 93.0 and 95.9%. The precision (coefficient of variance) of the method was in the range 0.8−12.3%. The method detection limit (MDL) and the method quantification limit (MQL) were 18.5 and 55.6 ng/g (ppb), respectively. The ergosterol analysis method developed can be used to handle 80 samples daily by one person, making it suitable for screening cereal cultivars for resistance to fungal infection. The ability for detecting low levels of ergosterol in a single kernel provides a tool to investigate early fungal invasion and to study mechanisms of resistance to fungal diseases.
Barley stem rust resistance genes: structure and function
(Plant Genome, 2009-07) Steffenson, Brian; Kleinhofs, Andris; Brueggeman, Robert; Nirmala, Jayaveeramuthu; Zhang, Ling; Mirlohi, Aghafakhr; Druka, Arnis; Rostoks, Nils
Rusts are biotrophic pathogens that attack many plant species but are particularly destructive on cereal crops. The stem rusts (caused by Puccinia graminis) have historically caused severe crop losses and continue to threaten production today. Barley (Hordeum vulgare L.) breeders have controlled major stem rust epidemics since the 1940s with a single durable resistance gene Rpg1. As new epidemics have threatened, additional resistance genes were identified to counter new rust races, such as the rpg4/Rpg5 complex locus against races QCCJ and TTKSK. To understand how these genes work, we initiated research to clone and characterize them. The Rpg1 gene encodes a unique protein kinase with dual kinase domains, an active kinase, and a pseudokinase. Function of both domains is essential to confer resistance. The rpg4 and Rpg5 genes are closely linked and function coordinately to confer resistance to several wheat (Triticum aestivum L.) stem rust races, including the race TTKSK (also called Ug99) that threatens the world's barley and wheat crops. The Rpg5 gene encodes typical resistance gene domains NBS, LRR, and protein kinase but is unique in that all three domains reside in a single gene, a previously unknown structure among plant disease resistance genes. The rpg4 gene encodes an actin depolymerizing factor that functions in cytoskeleton rearrangement.
The barley stem rust-resistance gene Rpg1 is a novel disease-resistance gene with homology to receptor kinases
(Proceedings of the National Academy of Sciences of the United States of America, 2002-07-09) Steffenson, Brian; Brueggeman, R.; Rostoks, N.; Kudrna, D.; Kilian, A.; Han, F.; Chen, J.; Druka, A.; Kleinhofs, A.
Stem rust caused by Puccinia graminis f. sp. tritici was among the most devastating diseases of barley in the northern Great Plains of the U.S. and Canada before the deployment of the stem rust-resistance gene Rpg1 in 1942. Since then, Rpg1 has provided durable protection against stem rust losses in widely grown barley cultivars (cvs.). Extensive efforts to clone Rpg1 by synteny with rice provided excellent flanking markers but failed to yield the gene because it does not seem to exist in rice. Here we report the map-based cloning and characterization of Rpg1. A high-resolution genetic map constructed with 8,518 gametes and a 330-kb bacterial artificial chromosome contig physical map positioned the gene between two crossovers ≈0.21 centimorgan and 110 kb apart. The region including Rpg1 was searched for potential candidate genes by sequencing low-copy probes. Two receptor kinase-like genes were identified. The candidate gene alleles were sequenced from resistant and susceptible cvs. Only one of the candidate genes showed a pattern of apparently functional gene structure in the resistant cvs. and defective gene structure in the susceptible cvs. identifying it as the Rpg1 gene. Rpg1 encodes a receptor kinase-like protein with two tandem protein kinase domains, a novel structure for a plant disease-resistance gene. Thus, it may represent a new class of plant resistance genes.
Changing the Game: Using Integrative Genomics to Probe Virulence Mechanisms of the Stem Rust Pathogen Puccinia graminis f. sp. tritici
(Frontiers in Plant Science, 2016-02-24) Steffenson, Brian; Figueroa, Melania; Upadhyaya, Narayana M; Sperschneider, Jana; Park, Robert F; Szabo, Les J; Ellis, Jeff G; Dodds, Peter N
The recent resurgence of wheat stem rust caused by new virulent races of Puccinia graminis f. sp. tritici (Pgt) poses a threat to food security. These concerns have catalyzed an extensive global effort toward controlling this disease. Substantial research and breeding programs target the identification and introduction of new stem rust resistance (Sr) genes in cultivars for genetic protection against the disease. Such resistance genes typically encode immune receptor proteins that recognize specific components of the pathogen, known as avirulence (Avr) proteins. A significant drawback to deploying cultivars with single Sr genes is that they are often overcome by evolution of the pathogen to escape recognition through alterations in Avr genes. Thus, a key element in achieving durable rust control is the deployment of multiple effective Sr genes in combination, either through conventional breeding or transgenic approaches, to minimize the risk of resistance breakdown. In this situation, evolution of pathogen virulence would require changes in multiple Avr genes in order to bypass recognition. However, choosing the optimal Sr gene combinations to deploy is a challenge that requires detailed knowledge of the pathogen Avr genes with which they interact and the virulence phenotypes of Pgt existing in nature. Identifying specific Avr genes from Pgt will provide screening tools to enhance pathogen virulence monitoring, assess heterozygosity and propensity for mutation in pathogen populations, and confirm individual Sr gene functions in crop varieties carrying multiple effective resistance genes. Toward this goal, much progress has been made in assembling a high quality reference genome sequence for Pgt, as well as a Pan-genome encompassing variation between multiple field isolates with diverse virulence spectra. In turn this has allowed prediction of Pgt effector gene candidates based on known features of Avr genes in other plant pathogens, including the related flax rust fungus. Upregulation of gene expression in haustoria and evidence for diversifying selection are two useful parameters to identify candidate Avr genes. Recently, we have also applied machine learning approaches to agnostically predict candidate effectors. Here, we review progress in stem rust pathogenomics and approaches currently underway to identify Avr genes recognized by wheat Sr genes.
Comparative Genome Structure, Secondary Metabolite, and Effector Coding Capacity across Cochliobolus Pathogens
(PLoS Genetics, 2013-01-24) Steffenson, Brian; Condon, Bradford J; Leng, Yueqiang; Wu, Dongliang; Bushley, Kathryn E; Ohm, Robin A; Otillar, Robert; Martin, Joel; Schackwitz, Wendy; Grimwood, Jane; MohdAzinudin, NurAinizzati; Xue, Chunsheng; Wang, Rui; Manning, Viola A; Dhillon, Braham; Tu, Zheng Jin; Salamov, Asaf; Sun, Hui; Lowry, Steve; LaButti, Kurt; Han, James; Copeland, Alex; Lindquist, Erika; Barry, Kerrie; Schmutz, Jeremy; Baker, Scott E; Ciuffetti, Lynda M; Grigoriev, Igor V; Zhong, Shaobin; Turgeon, B.Gillian
The genomes of five Cochliobolus heterostrophus strains, two Cochliobolus sativus strains, three additional Cochliobolus species (Cochliobolus victoriae, Cochliobolus carbonum, Cochliobolus miyabeanus), and closely related Setosphaeria turcica were sequenced at the Joint Genome Institute (JGI). The datasets were used to identify SNPs between strains and species, unique genomic regions, core secondary metabolism genes, and small secreted protein (SSP) candidate effector encoding genes with a view towards pinpointing structural elements and gene content associated with specificity of these closely related fungi to different cereal hosts. Whole-genome alignment shows that three to five percent of each genome differs between strains of the same species, while a quarter of each genome differs between species. On average, SNP counts among field isolates of the same C. heterostrophus species are more than 25× higher than those between inbred lines and 50× lower than SNPs between Cochliobolus species. The suites of nonribosomal peptide synthetase (NRPS), polyketide synthase (PKS), and SSP–encoding genes are astoundingly diverse among species but remarkably conserved among isolates of the same species, whether inbred or field strains, except for defining examples that map to unique genomic regions. Functional analysis of several strain-unique PKSs and NRPSs reveal a strong correlation with a role in virulence.
Development of a genetic linkage map for Sharon goatgrass (Aegilops sharonensis) and mapping of a leaf rust resistance gene
(Genome, 2013) Steffenson, Brian; Olivera, P.D.; Kilian, A.; Wenzl, P.
Aegilops sharonensis (Sharon goatgrass), a diploid wheat relative, is known to be a rich source of disease resistance genes for wheat improvement. To facilitate the transfer of these genes into wheat, information on their chromosomal location is important. A genetic linkage map of Ae. sharonensis was constructed based on 179 F2 plants derived from a cross between accessions resistant (1644) and susceptible (1193) to wheat leaf rust. The linkage map was based on 389 markers (377 Diversity Arrays Technology (DArT) and 12 simple sequence repeat (SSR) loci) and was comprised of 10 linkage groups, ranging from 2.3 to 124.6 cM. The total genetic length of the map was 818.0 cM, with an average interval distance between markers of 3.63 cM. Based on the chromosomal location of 115 markers previously mapped in wheat, the four linkage groups of A, B, C, and E were assigned to Ae. sharonensis (Ssh) and homoeologous wheat chromosomes 6, 1, 3, and 2. The single dominant gene (designated LrAeSh1644) conferring resistance to leaf rust race THBJ in accession 1644 was positioned on linkage group A (chromosome 6Ssh) and was flanked by DArT markers wpt-9881 (at 1.9 cM distal from the gene) and wpt-6925 (4.5 cM proximal). This study clearly demonstrates the utility of DArT for genotyping uncharacterized species and tagging resistance genes where pertinent genomic information is lacking.
Disease and insect resistance in cultivated barley accessions from the USDA National Small Grains Collection
(Crop Science, 2005) Steffenson, Brian; Bonman, J.Michael; Bockelman, Harold E.; Jackson, Lee F.
Cultivated barley (Hordeum vulgare subsp. vulgare L.) accessions from the USDA-ARS National Small Grains Collection (NSGC) have been tested systematically for the past 20 yr for disease and insect resistance. In this study, we analyzed the resistance to barley yellow dwarf (BYD), spot blotch (SB) caused by Cochliobolus sativus (Ito and Kuribayashi) Drechs. ex Dastur, net blotch (NB) caused by Pyrenophora teres f. teres Drechs., stripe rust (SR) caused by Puccinia striiformis Westend. f. sp. hordei, and Russian wheat aphid (RWA), Diuraphis noxia (Mordvilko), with respect to (i) geographic origin of resistant accessions, (ii) relationship to other NSGC descriptor data, and (iii) relationships among resistances. "Centers of concentration" for certain resistances were identified: eastern Africa for several diseases, western Turkey and the Caucasus for SR resistance, eastern Asia for adult plant resistance to NB, and south-central Asia for RWA resistance. Stripe rust resistance was also associated with accessions originating from high altitude in eastern Africa (Ethiopia). Various associations between resistances and grain descriptors, plant habit, and landrace status were also found. Forty-eight accessions showed multiple resistances on the basis of the field disease data and the RWA greenhouse data. Many of these resistant accessions were from Ethiopia, and many were of unknown origin. Stripe rust testing in California and Bolivia supported the conclusion that winter-habit accessions were more resistant to the disease than were spring-habit accessions. Information from this study will be used to guide future NSGC acquisition and evaluation efforts.
Effect of incubation time and temperature on the phenotypic expression of rpg4 to Puccinia graminis f. sp. tritici in barley
(Canadian Journal of Plant Pathology, 1997-03-01) Sun, Yonglian; Steffenson, Brian
To study the effect of incubation time and temperature on the phenotypic expression of rpg4, five barley genotypes with this resistance gene were infected with pathotype QCCJ of Puccinia graminis f. sp. tritici at the seedling stage, then subjected to various times of incubation at either 18-19°C or 27~28°C. Genotypes with rpg4 exhibited low (0, 0;, and 1), mesothetic (e.g. 3-210;, 120;3), and high (3,3) infection types at 18-19°C after initial incubation at 27-28°C for 0-28, 40-76, and 88 or more hours, respectively. A period of 88 or more hours of initial incubation at high temperature rendered the rpg4 resistance completely ineffective against this pathotype of P. g. f. sp. tritici. In contrast, high, mesothetic, and low infection types were found for the same genotypes at 27-28°C after initial incubation at 18-19°C for 0-40, 52-100, and 112 or more hours, respectively. The resistant infection types conferred by rpg4 are apparently established within the first 112 hours after the end of the infection period since subsequent shifts to higher temperature did not result in marked changes in the resistance response. These data indicate the critical importance of maintaining precise temperature control when assessing the infection phenotypes of barley genotypes carrying the stem rust resistance gene rpg4.
Genetic and molecular characterization of mating type genes in Cochliobolus sativus
(Mycologia, 2001-09-01) Steffenson, Brian; Zhong, Shaobin
Genetic and molecular approaches were used to characterize the mating type (MAT) genes in Cochliobolus sativus. One hundred and four ascospore progeny derived from a cross of C. sativus isolates ND93-1 (MAT-1) X ND9OPr (MAT-2) were backcrossed with their parents to determine mating type, but only five progeny produced pseudothecia with asci and/or ascospores. When degenerate primers from the conserved high mobility group (HMG) protein domain encoded by the MAT-2 gene in Cochliobolus species were used in polymerase chain reaction (PCR) with genomic DNA of C. sativus as templates, an amplicon of predicted size was amplified only from MAT-2 isolates. The presence of a MAT-2 homolog in these MAT-2 isolates was confirmed by Southern hybridization with the HMG box as a probe. Additionally, the presence or absence of the HMG homolog in the progeny segregated in a 1:1 ratio, as expected for the single gene control of mating type. Using primers based on the conserved regions at the 5' and 3' flanks of the idiomorphs in the MAT genes of other Cochliobolus species, the full-length MAT-1 and MAT-2 idiomorphs were cloned by PCR from C. sativus isolates ND93-1 and ND9OPr, respectively. DNA sequence analysis indicated that these two idiomorphs are organized in a manner similar to their respective counterparts in other Cochliobolus species. DNA hybridization and PCR amplification analysis of 54 field isolates of C. sativus collected worldwide showed that both mating types exist in populations round the world. The low frequency of successful backcrosses of progeny to parents in the ND93-1 X ND9OPr cross, combined with the fact that many crosses between isolates of opposite mating type are unsuccessful, suggests that genetic factors other than MAT genes affect the fertility of the fungus.
Genetic architecture of quantitative trait loci associated with morphological and agronomic trait differences in a wild by cultivated barley cross
(Genome, 2007) Steffenson, Brian; Gyenis, L.; Yun, S.J.; Smith, K.P.; Bossolini, E.; Sanguineti, M.C.; Muehlbauer, G.J.
Hordeum vulgare subsp. spontaneum is the progenitor of cultivated barley (Hordeum vulgare L.). Domestication combined with plant breeding has led to the morphological and agronomic characteristics of modern barley cultivars. The objective of this study was to map the genetic factors that morphologically and agronomically differentiate wild barley from modern barley cultivars. To address this objective, we identified quantitative trait loci (QTLs) associated with plant height, flag leaf width, spike length, spike width, glume length in relation to seed length, awn length, fragility of ear rachis, endosperm width and groove depth, heading date, flag leaf length, number of tillers per plant, and kernel color in a Harrington/OUH602 advanced backcross (BC2F8) population. This population was genotyped with 113 simple sequence repeat markers. Thirty QTLs were identified, of which 16 were newly identified in this study. One to 4 QTLs were identified for each of the traits except glume length, for which no QTL was detected. The portion of phenotypic variation accounted for by individual QTLs ranged from about 9% to 54%. For traits with more than one QTL, the phenotypic variation explained ranged from 25% to 71%. Taken together, our results reveal the genetic architecture of morphological and agronomic traits that differentiate wild from cultivated barley.
Genetic variation for virulence and RFLP markers in Pyrenophora teres
(Canadian Journal of Plant Pathology, 2003-03-01) Wu, H.-L.; Steffenson, Brian; Zhong, S; Li, Y; Oleson, A.E.
Pyrenophora teres f. teres (causing net blotch) and Pyrenophora teres f. maculata (causing "spot form" of the disease) are important foliar pathogens of barley. In breeding for resistance to disease, it is important to have a thorough knowledge of the degree of genetic variation in the pathogen. This study was undertaken to assess genetic variation in a small, but geographically diverse collection of P. teres isolates. Isolates derived from single conidia were evaluated for their virulence phenotypes on 25 differential barley genotypes. Fifteen pathotypes were identified from a collection of 23 P. t. f. teres isolates, and 4 pathotypes, from a collection of 8 P. t. f. maculata isolates. In general, the P. t. f. teres isolates exhibited a broader spectrum and a higher level of virulence on the host differentials than the P. t. f. maculata isolates. Eight barley genotypes were resistant to all 19 pathotypes identified and should be useful in breeding barley for resistance to both forms of P. teres. Genetic variation was also examined by restriction fragment length polymorphism (RFLP) analysis. A 0.46-kb DNA fragment (ND218) generated by the polymerase chain reaction from genomic DNA of a California isolate of P. t. f. teres was used as a probe. Every P. teres isolate tested with ND218 exhibited a unique RFLP pattern. Cluster analysis, based on both the virulence phenotypes and RFLP patterns, indicates that P. teres possesses a high degree of diversity at the species and subspecies levels. The high degree of polymorphism revealed by ND218 will make this probe a useful tool for the DNA fingerprinting of P. teres isolates.
Genetically engineered stem rust resistance in barley using the Rpg1 gene
(Proceedings of the National Academy of Sciences of the United States of America, 2003-01-07) Steffenson, Brian; Horvath, Henriette; Rostoks, Nils; Brueggeman, Robert; Wettstein, Diter von; Kleinhofs, Andris
The stem-rust-susceptible barley cv. Golden Promise was transformed by Agrobacterium-mediated transformation of immature zygotic embryos with the Rpg1 genomic clone of cv. Morex containing a 520-bp 5′ promoter region, 4,919-bp gene region, and 547-bp 3′ nontranscribed sequence. Representatives of 42 transgenic barley lines obtained were characterized for their seedling infection response to pathotype Pgt-MCC of the stem rust fungus Puccinia graminis f. sp. tritici. Golden Promise was converted from a highly susceptible cultivar into a highly resistant one by transformation with the dominant Rpg1 gene. A single copy of the gene was sufficient to confer resistance against stem rust, and progenies from several transformants segregated in a 3:1 ratio for resistance/susceptibility as expected for Mendelian inheritance. These results unequivocally demonstrate that the DNA segment isolated by map-based cloning is the functional Rpg1 gene for stem rust, resistance. One of the remarkable aspects about the transformants is that they exhibit a higher level of resistance than the original sources of Rpg1 (cvs. Chevron and Peatland). In most cases, the Golden Promise transformants exhibited a highly resistant reaction where no visible sign of infection was evident. Hypersensitive necrotic “fleck” reactions were also observed, but less frequently. With both infection types, pathogen sporulation was prevented. Southern blot and RT-PCR analysis revealed that neither Rpg1 gene copy number nor expression levels could account for the increased resistance observed in Golden Promise transformants. Nevertheless, this research demonstrates that stem-rust-susceptible barley can be made resistant by transformation with the cloned Rpg1 gene.
Genome-wide association mapping of fusarium head blight resistance and agromorphological traits in barley landraces from Ethiopia and Eritrea
(Crop Science, 2015-06-12) Steffenson, Brian; Mamo, Bullo Erena
Fusarium head blight (FHB), caused primarily by Fusarium graminearum, is an important disease of barley (Hordeum vulgare L.), and other cereals. In barley, the genetic basis of FHB resistance has been intensively studied through linkage mapping that identified several quantitative trait loci (QTL). However, our understanding and application of these QTL in breeding is still limited due to the complex nature and low-to-moderate heritability of FHB resistance. Previous studies used either breeding lines, unimproved varieties, or germplasm selections. Here, we used association mapping in barley landraces to identify QTL associated with FHB severity, deoxynivalenol (DON) concentration and correlated agromorphological traits. Diverse barley landraces (n = 298) from Ethiopia and Eritrea were evaluated for the traits under field conditions for 2 yr (2011–2012) in Crookston, MN, and genotyped with 7842 single nucleotide polymorphism (SNP) markers. Association mapping analysis using a mixed model corrected for pairwise relatedness between individuals identified one common resistance QTL on barley chromosome 2HL significantly associated with both FHB severity and DON concentration and another one on 4HL associated with DON concentration. The QTL identified on 2HL is associated with the row-type locus Vrs1. Both of these QTL were not significantly associated with heading date or plant height unlike other QTL reported in previous studies. Thus, the resistant accessions carrying these QTL may be used in breeding programs without the confounding effects from these agromorphological traits. Importantly, these QTL could be new alleles preserved in this unique germplasm, and the linked SNP markers found may be useful in marker-assisted introgression of resistance.
A genome-wide association study for culm cellulose content in barley reveals candidate genes co-expressed with members of the cellulose synthase a gene family
(PLoS ONE, 2015-07-08) Steffenson, Brian; Houston, Kelly; Burton, Rachel A; Sznajder, Beata; Rafalski, Antoni J; Dhugga, Kanwarpal S; Mather, Diane E; Taylor, Jillian; Waugh, Robbie; Fincher, Geoffrey B
Cellulose is a fundamentally important component of cell walls of higher plants. It provides a scaffold that allows the development and growth of the plant to occur in an ordered fashion. Cellulose also provides mechanical strength, which is crucial for both normal development and to enable the plant to withstand both abiotic and biotic stresses. We quantified the cellulose concentration in the culm of 288 two – rowed and 288 six – rowed spring type barley accessions that were part of the USDA funded barley Coordinated Agricultural Project (CAP) program in the USA. When the population structure of these accessions was analysed we identified six distinct populations, four of which we considered to be comprised of a sufficient number of accessions to be suitable for genome-wide association studies (GWAS). These lines had been genotyped with 3072 SNPs so we combined the trait and genetic data to carry out GWAS. The analysis allowed us to identify regions of the genome containing significant associations between molecular markers and cellulose concentration data, including one region cross-validated in multiple populations. To identify candidate genes we assembled the gene content of these regions and used these to query a comprehensive RNA-seq based gene expression atlas. This provided us with gene annotations and associated expression data across multiple tissues, which allowed us to formulate a supported list of candidate genes that regulate cellulose biosynthesis. Several regions identified by our analysis contain genes that are co-expressed with CELLULOSE SYNTHASE A (HvCesA) across a range of tissues and developmental stages. These genes are involved in both primary and secondary cell wall development. In addition, genes that have been previously linked with cellulose synthesis by biochemical methods, such as HvCOBRA, a gene of unknown function, were also associated with cellulose levels in the association panel. Our analyses provide new insights into the genes that contribute to cellulose content in cereal culms and to a greater understanding of the interactions between them.
Identification and chromosomal location of major genes for resistance to Pyrenophora teres in a doubled-haploid barley population
(Genome, 2006) Steffenson, Brian; Friesen, T.L.; Faris, J.D.; Lai, Z.
Net blotch, caused by Pyrenophora teres, is one of the most economically important diseases of barley worldwide. Here, we used a barley doubled-haploid population derived from the lines SM89010 and Q21861 to identify major quantitative trait loci (QTLs) associated with seedling resistance to P. teres f. teres (net-type net blotch (NTNB)) and P. teres f. maculata (spot-type net blotch (STNB)). A map consisting of simple sequence repeat (SSR) and amplified fragment length polymorphism (AFLP) markers was used to identify chromosome locations of resistance loci. Major QTLs for NTNB and STNB resistance were located on chromosomes 6H and 4H, respectively. The 6H locus (NTNB) accounted for as much as 89% of the disease variation, whereas the 4H locus (STNB resistance) accounted for 64%. The markers closely linked to the resistance gene loci will be useful for marker-assisted selection.Key words: disease resistance, Drechslera teres, molecular markers.
Identification and mapping of a leaf rust resistance gene in barley line Q21861
(Genome, 1997) Steffenson, Brian; Borovkova, I.G.; Jin, Y.; Kilian, A.; Blake, T.K.; Kleinhofs, A.
Barley line Q21861 possesses an incompletely dominant gene (RphQ) for resistance to leaf rust caused by Puccinia hordei. To investigate the allelic and linkage relations between RphQ and other known Rph genes, F2 populations from crosses between Q21861 and donors of Rph1 to Rph14 (except for Rph8) were evaluated for leaf rust reaction at the seedling stage. Results indicate that RphQ is either allelic with or closely linked to the Rph2 locus. A doubled haploid population derived from a cross between Q21861 and SM89010 (a leaf rust susceptible line) was used for molecular mapping of the resistance locus. Bulked segregant analysis was used to identify markers linked to RphQ, using random amplified polymorphic DNAs (RAPDs), restriction fragment length polymorphisms (RFLPs), and sequence tagged sites (STSs). Of 600 decamer primers screened, amplified fragments generated by 9 primers were found to be linked to the RphQ locus; however, only 4 of them were within 10 cM of the target. The RphQ locus was mapped to the centromeric region of chromosome 7, with a linkage distance of 3.5 cM from the RFLP marker CDO749. Rrn2, an RFLP clone from the ribosomal RNA intergenic spacer region, was found to be very closely linked with RphQ, based on bulked segregant analysis. An STS marker, ITS1, derived from Rrn2, was also closely linked (1.6 cM) to RphQ.
Identification of Cochliobolus sativus isolates expressing differential virulence on two-rowed barley genotypes from North Dakota
(Canadian Journal of Plant Pathology, 2009-12-29) Fetch Jr, T.G.; Steffenson, Brian
Severe spot blotch infection was observed in 1990 on several two-row barley breeding lines previously regarded as resistant to Cochliobolus sativus. Studies were conducted to compare the virulence pattern of a C. sativus isolate (ND90Pr) obtained from this two-row breeding nursery with one (ND85F) used in previous disease screening evaluations. Greenhouse and field experiments were performed in 1991 and 1992 at Fargo, ND, using a split plot design with isolate as the main effect. Isolates ND90Pr and ND85F exhibited distinct differential virulence patterns on barley genotypes ND 5883, ND 12437, ND 12720, ND 12721, and Bowman. Isolate ND90Pr displayed high virulence on ND 12720, ND 12721, and Bowman, and low virulence on ND 5883 and ND 12437. In contrast, isolate ND85F was highly virulent on ND 5883 and ND 12437 and weakly virulent on ND 12720, ND 12721, and Bowman. Both isolates expressed low virulence on genotype ND Bl 12, the primary source of resistance to C. sativus in commercial six-row barley germplasm. To incorporate adequate levels of resistance into future two-row barley cultivars, disease evaluations should be made with C. sativus isolates that express the full spectrum of virulence found in North Dakota.

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