Identification and genetic mapping of a recessive gene for resistance to stripe rust in wheat line LM168-1

Stripe rust (or yellow rust), caused by the fungus Puccinia striiformis f. sp. tritici (Pst), is one of the most important foliar diseases of wheat. Characterization and utilization of novel resistant genes is the most effective, economic and environmentally friendly approach to controlling the disease. Wheat line LM168-1, which was derived from a cross between common wheat Chuannong 16 and Milan, has good adult-plant resistance to stripe rust, based on field tests over several years. To elucidate the genetic basis of resistance, LM168-1 was crossed with susceptible variety SY95-71. Parents and F1, F2, BC1 and F2:3 progenies were tested in 2009–2011 in a field inoculated with the predominant races of Pst in China. The genetic analysis showed that resistance to stripe rust in LM168-1 was controlled by a single recessive gene, temporarily designated yrLM168. Simple sequence repeat (SSR), resistance gene analog polymorphism (RGAP) and target region amplification polymorphism (TRAP) techniques were used to identify molecular markers linked to the resistance locus. Finally, a linkage group consisting of two SSR, four RGAP and five TRAP markers was constructed for yrLM168 with 102 F2 plants. The closest markers R1 and R2 flanked the resistance gene locus at 2.4 and 2.4 cM, respectively. Furthermore, two SSR markers Xwmc59 and Xwmc145 assigned the gene to chromosome 6A. Because yrLM168 confers high-level resistance to the predominant races of Pst in China, it should be useful in stripe rust resistance breeding programs. The closely linked markers can be used for rapidly transferring yrLM168 to wheat breeding populations.     

Genetics and mapping of stem rust resistance to Ug99 in the wheat cultivar Webster

New races of wheat stem rust, namely TTKSK (Ug99) and its variants, pose a threat to wheat production in the regions where they are found. The accession of the wheat cultivar Webster (RL6201) maintained at the Cereal Research Centre in Winnipeg, Canada, shows resistance to TTKSK and other races of stem rust. The purpose of this study was to study the inheritance of seedling resistance to stem rust in RL6201 and genetically map the resistance genes using microsatellite (SSR) markers. A population was produced by crossing the stem rust susceptible line RL6071 with Webster. The F₂ and F₃ were tested with TPMK, a stem rust race native to North America. The F₃ was also tested with TTKSK. Two independently assorting genes were identified in RL6201. Resistance to TPMK was conferred by Sr30, which was mapped with microsatellites on chromosome 5DL. The second gene, temporarily designated SrWeb, conferred resistance to TTKSK. SrWeb was mapped to chromosome 2BL using SSR markers. Comparison with previous genetic maps showed that SrWeb occupies a locus near Sr9. Further analysis will be required to determine if SrWeb is a new gene or an allele of a previously identified gene.     

Overexpression of a western white pine PR10 protein enhances cold tolerance in transgenic Arabidopsis

The PmPR10-1.10 protein from western white pine is known to be associated with frost hardiness, and up-regulated by seasonal cold acclimation and biotic and abiotic stresses. To gain insight into the molecular basis of cold hardiness, we investigated the potential physiological role of PmPR10-1.10 by gene overexpression in transgenic Arabidopsis plants. A binary vector was constructed for PmPR10-1.10 synthesis in higher plants and transgenic Arabidopsis lines were generated by Agrobacterium-mediated transformation. Following Western protein blot analysis confirming target protein production, transgenic Arabidopsis lines were tested for cold tolerance by electrolyte leakage analysis post treatment of different freezing temperatures. Our results demonstrate that accumulation of PmPR10-1.10 protein resulted in significantly greater freezing tolerance in transgenic plants than in wild type plants. This indicates that the transfer and selection of cold acclimation proteins like PmPR10-1.10 may be a breeding strategy for the development of freezing tolerance in conifers.     

Genetic analysis and molecular mapping of a stripe rust resistance gene derived from Psathynrostachys huashanica Keng in wheat line H9014-121-5-5-9

Stripe rust, caused by Puccinia striiformis f. sp. tritici (Pst), is one of the most devastating diseases worldwide and is also an important disease in China. The wheat translocation line H9014-121-5-5-9 was originally developed from interspecific hybridization between wheat (Triticum aestivum L.) line 7182 and Psathyrostachys huashanica Keng. This translocation line showed resistance to predominant stripe rust races in China when it was tested with nine races of Pst. To determine the inheritance and map the resistance gene, segregating populations were developed from the cross between H9014-121-5-5-9 and the susceptible cultivar Mingxian 169. The seedlings of the F1, F2, and F2:3 generations were tested with race CYR31. The results showed that the resistance in H9014-121-5-5-9 was conferred by a single dominant gene. Bulked segregant analysis and simple sequence repeat (SSR) markers were used to identify polymorphic markers associated with the resistance gene locus. Seven polymorphic SSR markers were linked to the resistance gene. A linkage map was constructed according to the genotypes of the seven SSR markers and the resistance gene. Based on the SSR marker positions on the wheat chromosome, the resistance gene was assigned on chromosome 1AL, temporarily designated YrHA. Based on chromosomal location, reaction patterns and pedigree analysis, YrHA should be a novel resistance gene to stripe rust. The molecular markers of the new resistance gene in H9014-121-5-5-9 could be useful for marker-assisted selection in breeding programs against stripe rust.     

Simultaneous transfer, introgression, and genomic localization of genes for resistance to stem rust race TTKSK (Ug99) from Aegilops tauschii to wheat

Wheat production is currently threatened by widely virulent races of the wheat stem rust fungus, Puccinia graminis f. sp. tritici, that are part of the TTKSK (also known as ‘Ug99’) race group. The diploid D genome donor species Aegilops tauschii (2n = 2x = 14, DD) is a readily accessible source of resistance to TTKSK and its derivatives that can be transferred to hexaploid wheat, Triticum aestivum (2n = 6x = 42, AABBDD). To expedite transfer of TTKSK resistance from Ae. tauschii, a direct hybridization approach was undertaken that integrates gene transfer, mapping, and introgression into one process. Direct crossing of Ae. tauschii accessions with an elite wheat breeding line combines the steps of gene transfer and introgression while development of mapping populations during gene transfer enables the identification of closely linked markers. Direct crosses were made using TTKSK-resistant Ae. tauschii accessions TA1662 and PI 603225 as males and a stem rust-susceptible T. aestivum breeding line, KS05HW14, as a female. Embryo rescue enabled recovery of F₁ (ABDD) plants that were backcrossed as females to the hexaploid recurrent parent. Stem rust-resistant BC₁F₁ plants from each Ae. tauschii donor source were used as males to generate BC₂F₁ mapping populations. Bulked segregant analysis of BC₂F₁ genotypes was performed using 70 SSR loci distributed across the D genome. Using this approach, stem rust resistance genes from both accessions were located on chromosome arm 1DS and mapped using SSR and EST-STS markers. An allelism test indicated the stem rust resistance gene transferred from PI 603225 is Sr33. Race specificity suggests the stem rust resistance gene transferred from TA1662 is unique and this gene has been temporarily designated SrTA1662. Stem rust resistance genes derived from TA1662 and PI 603225 have been made available with selectable molecular markers in genetic backgrounds suitable for stem rust resistance breeding.     

Chromosome location, DNA markers and rust resistance of the sunflower gene R 5

Sunflower rust, caused by the fungus Puccinia helianthi Schwein., was not a serious problem for many decades because of successful deployment of effective resistance genes in commercial sunflower (Helianthus annuus L.) hybrids in North America. In the 1980s and early 1990s, however, a shift in virulence of the rust race population in North America rendered most of the commercial hybrids susceptible to new virulent races. A germplasm line, HA-R2, carrying the rust resistance gene R ₅ was released as a multi-race rust-resistant line in 1985 but has not been widely used in commercial hybrid production. R ₅ remains effective against the prevalent rust races of sunflower in North America. This gene was previously reported to be associated with two simple sequence repeat (SSR) markers, ORS316 and ORS630, which were mapped to linkage group (LG) 13 of sunflower. However, out of the 63 markers of LG13 screened in the present study, only 18, including ORS316 and ORS630, were polymorphic. These markers, which covered all of LG 13, were assayed in 94 individual F2 progenies derived from the cross of HA 89 with HA-R2. All failed to detect any locus in LG13 associated with the gene R ₅ . Subsequently, a bulked segregant analysis was employed with an additional 510 SSR markers selected from the remaining 16 LGs of the sunflower genome. This analysis demonstrated that the LG2 markers showed association with rust resistance. Genotyping of the 94 F2 individuals with 23 polymorphic SSR markers from LG2 confirmed the R ₅ location on LG2, flanked by two SSR markers, ORS1197-2 and ORS653a, at 3.3 and 1.8?cM of genetic distance, respectively. The markers for R ₅ developed in this study will provide a useful tool for speeding up deployment of the R ₅ gene in commercial sunflower hybrid production.     

Development and characterization of wheat lines carrying stem rust resistance gene Sr43 derived from Thinopyrum ponticum

Key message

Wheat lines carrying Ug99-effective stem rust resistance gene Sr43 on shortened alien chromosome segments were produced using chromosome engineering, and molecular markers linked to Sr43 were identified for marker-assisted selection.

Abstract

Stem rust resistance gene Sr43, transferred into common wheat (Triticum aestivum) from Thinopyrum ponticum, is an effective gene against stem rust Ug99 races. However, this gene has not been used in wheat breeding because it is located on a large Th. ponticum 7el₂ chromosome segment, which also harbors genes for undesirable traits. The objective of this study was to eliminate excessive Th. ponticum chromatin surrounding Sr43 to make it usable in wheat breeding. The two original translocation lines KS10-2 and KS24-1 carrying Sr43 were first analyzed using simple sequence repeat (SSR) markers and florescent genomic in situ hybridization. Six SSR markers located on wheat chromosome arm 7DL were identified to be associated with the Th. ponticum chromatin in KS10-2 and KS24-1. The results confirmed that KS24-1 is a 7DS·7el₂L Robertsonian translocation as previously reported. However, KS10-2, which was previously designated as a 7el₂S·7el₂L-7DL translocation, was identified as a 7DS-7el₂S·7el₂L translocation. To reduce the Th. ponticum chromatin carrying Sr43, a BC₂F₁ population (Chinese Spring//Chinese Spring ph1bph1b*2/KS10-2) containing ph1b-induced homoeologous recombinants was developed, tested with stem rust, and genotyped with the six SSR markers identified above. Two new wheat lines (RWG33 and RWG34) carrying Sr43 on shortened alien chromosome segments (about 17.5 and 13.7 % of the translocation chromosomes, respectively) were obtained, and two molecular markers linked to Sr43 in these lines were identified. The new wheat lines with Sr43 and the closely linked markers provide new resources for improving resistance to Ug99 and other races of stem rust in wheat.     

Molecular mapping of leaf rust resistance gene LrNJ97 in Chinese wheat line Neijiang 977671

Neijiang 977671 and 19 near-isogenic lines with known leaf rust resistance genes were inoculated with 12 pathotypes of Puccinia triticina for postulation of leaf rust resistance genes effective at the seedling stage. The reaction pattern of Neijiang 977671 differed from those of the lines with known leaf rust resistance genes used in the test, indicating that Neijiang 977671 may carry a new leaf rust resistance gene(s). With the objective of identifying and mapping the new gene for resistance to leaf rust, F₁ and F₂ plants, and F_2:3 families, from Neijiang 977671 × Zhengzhou 5389 (susceptible) were inoculated with Chinese P. triticina pathotype FHNQ in the greenhouse. Results from the F₂ and F_2:3 populations indicated that a single dominant gene, temporarily designated LrNJ97, conferred resistance. In order to identify other possible genes in Neijiang 977671 other eight P. triticina pathotypes avirulent on Neijiang 977671 were used to inoculate 25 F_2:3 families. The results showed that at least three leaf rust resistance genes were deduced in Neijiang 977671. Bulked segregant analysis was performed on equal amounts of genomic DNA from 20 resistant and 20 susceptible F₂ plants. SSR markers polymorphic between the resistant and susceptible bulks were used to analyze the F_2:3 families. LrNJ97 was linked to five SSR loci on chromosome 2BL. The two closest flanking SSR loci were Xwmc317 and Xbarc159 at genetic distances of 4.2 and 2.2 cM, respectively. At present two designated genes (Lr50 and Lr58) are located on chromosome 2BL. In the seedling tests, the reaction pattern of LrNJ97 was different from that of Lr50. Lr50 and Lr58 were derived from T. armeniacum and Ae. triuncialis, respectively, whereas according to the pedigree of Neijiang 977671 LrNJ97 is from common wheat. Although seeds of lines with Lr58 were not available, it was concluded that LrNJ97 is likely to be a new leaf rust resistance gene.     

Mapping of stripe rust resistance gene in an <Emphasis Type="Italic">Aegilops caudata</Emphasis> introgression line in wheat and its genetic association with leaf rust resistance

A pair of stripe rust and leaf rust resistance genes was introgressed from Aegilops caudata, a nonprogenitor diploid species with the CC genome, to cultivated wheat. Inheritance and genetic mapping of stripe rust resistance gene in backcross-recombinant inbred line (BC-RIL) population derived from the cross of a wheat–Ae. caudata introgression line (IL) T291-2(pau16060) with wheat cv. PBW343 is reported here. Segregation of BC-RILs for stripe rust resistance depicted a single major gene conditioning adult plant resistance (APR) with stripe rust reaction varying from TR-20MS in resistant RILs signifying the presence of some minor genes as well. Genetic association with leaf rust resistance revealed that two genes are located at a recombination distance of 13%. IL T291-2 had earlier been reported to carry introgressions on wheat chromosomes 2D, 3D, 4D, 5D, 6D and 7D. Genetic mapping indicated the introgression of stripe rust resistance gene on wheat chromosome 5DS in the region carrying leaf rust resistance gene LrAc, but as an independent introgression. Simple sequence repeat (SSR) and sequence-tagged site (STS) markers designed from the survey sequence data of 5DS enriched the target region harbouring stripe and leaf rust resistance genes. Stripe rust resistance locus, temporarily designated as YrAc, mapped at the distal most end of 5DS linked with a group of four colocated SSRs and two resistance gene analogue (RGA)-STS markers at a distance of 5.3 cM. LrAc mapped at a distance of 9.0 cM from the YrAc and at 2.8 cM from RGA-STS marker Ta5DS_2737450, YrAc and LrAc appear to be the candidate genes for marker-assisted enrichment of the wheat gene pool for rust resistance.     

Characterization of Sr9h,a wheat stem rust resistance allele effective to Ug99

Key message

Wheat stem rust resistance gene SrWeb is an allele at the Sr9 locus that confers resistance to Ug99.

Abstract

Race TTKSK (Ug99) of Puccinia graminis f. sp. tritici, the causal fungus of stem rust, threatens global wheat production because of its broad virulence to current wheat cultivars. A recently identified Ug99 resistance gene from cultivar Webster, temporarily designated as SrWeb, mapped near the stem rust resistance gene locus Sr9. We determined that SrWeb is also present in Ug99 resistant cultivar Gabo 56 by comparative mapping and an allelism test. Analysis of resistance in a population segregating for both Sr9e and SrWeb demonstrated that SrWeb is an allele at the Sr9 locus, which subsequently was designated as Sr9h. Webster and Gabo 56 were susceptible to the Ug99-related race TTKSF+ from South Africa. Race TTKSF+ possesses unique virulence to uncharacterized Ug99 resistance in cultivar Matlabas. This result validated that resistance to Ug99 in Webster and Gabo 56 is conferred by the same gene: Sr9h. The emergence of pathogen virulence to several resistance genes that are effective to the original Ug99 race TTKSK, including Sr9h, suggests that resistance genes should be used in combinations in order to increase resistance durability.     

Resistance and virulence interactions between two white pine species and blister rust in a 30-year field trial

We monitored infection and mortality of full-sib families of sugar pine (SP) and western white pine (WWP) selected for different mechanisms of resistance to white pine blister rust for more than 30 years in a field test in northern California. Natural infection was enhanced by interplanting alternate host Ribes spp. among test seedlings. Parents of the families were from three geographic provinces, representing the northern Rocky Mountains of Idaho (WWP), the Cascade Range of Oregon and Washington (WWP), and the Siskiyou Mountains of California and Oregon (SP). Several families of each pine species had major gene resistance (MGR), or genes for partial resistance (PR), or both types. Under the chronic epidemic conditions that prevailed, susceptible controls of both species became infected and died within a few years. Families with MGR segregated in expected Mendelian ratios (susceptibility/resistance), but each R gene—Cr1 in sugar pine and Cr2 in western white pine—was eventually defeated by biotypes of the rust with specific virulence to it (vcr1 and vcr2, respectively). WWP families from Idaho, which lacked Cr2, performed the best overall, ranging from 44 to 64% rust-free; a few WWP families from the Cascades, both with and without Cr2, performed comparably. Families with PR were not specifically vulnerable to vcr1 or vcr2. PR was less frequent in SP than WWP, and SP families had steeper infection rate curves than WWP. Combining PR in pedigrees with MGR may prolong protection of MGR by inhibiting selection and epidemic increase of vcr genes in the rust.     

Identification and mapping in spring wheat of genetic factors controlling stem rust resistance and the study of their epistatic interactions across multiple environments

Stem rust (Puccinia graminis f. sp. tritici) is responsible for major production losses in hexaploid wheat (Triticum aestivum L.) around the world. The spread of stem rust race Ug99 and variants is a threat to worldwide wheat production and efforts are ongoing to identify and incorporate resistance. The objectives of this research were to identify quantitative trait loci (QTL) and to study their epistatic interactions for stem rust resistance in a population derived from the Canadian wheat cultivars AC Cadillac and Carberry. A doubled haploid (DH) population was developed and genotyped with DArT^® and SSR markers. The parents and DH lines were phenotyped for stem rust severity and infection response to Ug99 and variant races in 2009, 2010 and 2011 in field rust nurseries near Njoro, Kenya, and to North American races in 2011 and 2012 near Swift Current, SK, Canada. Seedling infection type to race TTKSK was assessed in a bio-containment facility in 2009 and 2012 near Morden, MB. Eight QTL for stem rust resistance and three QTL for pseudo-black chaff on nine wheat chromosomes were identified. The phenotypic variance (PV) explained by the stem rust resistance QTL ranged from 2.4 to 48.8 %. AC Cadillac contributed stem rust resistance QTL on chromosomes 2B, 3B, 5B, 6D, 7B and 7D. Carberry contributed resistance QTL on 4B and 5A. Epistatic interactions were observed between loci on 4B and 5B, 4B and 7B, 6D and 3B, 6D and 5B, and 6D and 7B. The stem rust resistance locus on 6D interacted synergistically with 5B to improve the disease resistance through both crossover and non-crossover interactions depending on the environment. Results from this study will assist in planning breeding for stem rust resistance by maximizing QTL main effects and epistatic interactions.     

QTL mapping of resistance to race Ug99 of Puccinia graminis f. sp. tritici in durum wheat (Triticum durum Desf.)

Stem rust caused by Puccinia graminis f. sp. tritici was historically one of the most destructive diseases of wheat worldwide. The evolution and rapid migration of race TTKSK (Ug99) and derivatives, first detected in Uganda in 1999, are of international concern due to the virulence of these races to widely used stem rust resistance genes. In attempts to identify quantitative trait loci (QTL) linked with resistance to stem rust race Ug99, 95 recombinant inbred lines that were developed from a cross between two durum wheat varieties, Kristal and Sebatel, were evaluated for reaction to stem rust. Seven field trials at two locations were carried out in main and off seasons. In addition to the natural infection, the nursery was also artificially inoculated with urediniospores of stem rust race Ug99 and a mixture of locally collected stem rust urediniospores. A genetic map was constructed based on 207 simple sequence repeat (SSR) and two sequence tagged site loci. Using composite interval mapping, nine QTL for resistance to stem rust were identified on chromosomes 1AL, 2AS, 3BS, 4BL, 5BL, 6AL 7A, 7AL and 7BL. These results suggest that durum wheat resistance to stem rust is oligogenic and that there is potential to identify previously uncharacterized resistance genes with minor effects. The SSR markers that are closely linked to the QTL can be used for marker-assisted selection for stem rust resistance in durum wheat.     

Development of leucine-rich repeat polymorphism, amplified fragment length polymorphism, and sequence characterized amplified region markers to the Cronartium ribicola resistance gene Cr2 in western white pine ( Pinus monticola )

White pine blister rust (WPBR), caused by Cronartium ribicola, is a devastating disease in Pinus monticola and other five-needle pines. Pyramiding a major resistance gene (Cr2) with other resistance genes is an important component of integrated strategies to control WPBR in P. monticola. To facilitate this strategy, the objective of the present study was to identify leucine-rich repeat (LRR) polymorphisms, amplified fragment length polymorphisms (AFLPs), and sequence characterized amplified region (SCAR) markers linked to the western white pine Cr2 (BSA) gene for precise gene mapping. Bulked segregant analysis and haploid segregation analysis allowed the identification of 11 LRR polymorphisms and five AFLP markers in the Cr2 linkage. The closest LRR markers were 0.53 Kosambi cM from Cr2 at either end. After marker cloning and sequencing, AFLP marker EacccMccgat-365 and random polymorphic DNA marker U570–843 were converted successfully into SCAR markers. For a potential application in marker-assisted selection (MAS), these two SCAR markers were verified in two western white pine families. This study represents the first report of LRR-related DNA markers linked to C. ribicola resistance in five-needle pines. These findings may help further candidate gene identification for disease resistance in a conifer species.     

Molecular mapping of a gene for stripe rust resistance in spring wheat cultivar IDO377s

Stripe rust, caused by Puccinia striiformis f. sp. tritici, is one of the most important diseases of wheat worldwide. The best strategy to control stripe rust is to grow resistant cultivars. One such cultivar resistant to most races in North America is ‘IDO377s’. To study the genetics of its resistance this spring wheat cultivar was crossed with ‘Avocet Susceptible’ (AvS). Seedlings of the parents, F₂ plants, and F₃ lines were tested under controlled greenhouse conditions with races PST-43 and PST-45 of P. striiformis f. sp. tritici. IDO377s carries a single dominant gene for resistance. Resistance gene analog polymorphism (RGAP) and simple sequence repeat (SSR) techniques were used to identify molecular markers linked to the resistance gene. A total of ten markers were identified, two of which flanked the locus at 4.4 and 5.5 cM. These flanking RGAP markers were located on chromosome 2B with nulli-tetrasomic lines of ‘Chinese Spring’. Their presence in the ditelosomic 2BL line localized them to the long arm. The chromosomal location of the resistance gene was further confirmed with two 2BL-specific SSR markers and a sequence tagged site (STS) marker previously mapped to 2BL. Based on the chromosomal location, reactions to various races of the pathogen and tests of allelism, the IDO377s gene is different from all previously designated genes for stripe rust resistance, and is therefore designated Yr43. A total of 108 wheat breeding lines and cultivars with IDO377s or related cultivars in their parentage were assayed to assess the status of the closest flanking markers and to select lines carrying Yr43. The results showed that the flanking markers were reliable for assisting selection of breeding lines carrying the resistance gene. A linked stripe rust resistance gene, previously identified as YrZak, in cultivar Zak was designated Yr44.     

Genetic dissection of fusiform rust and pitch canker disease traits in loblolly pine

Loblolly pine (Pinus taeda L.) exhibits genetic resistance to fusiform rust disease (incited by the biotrophic fungus, Cronartium quercuum f. sp. fusiforme) and pitch canker disease (incited by the necrotrophic fungus, Fusarium circinatum). In this study, a total of 14,015 loblolly pine cuttings from 1,065 clones were screened in controlled greenhouse conditions to identify phenotypes of clones, families, and parents that guide a genetic dissection of disease traits associated with pitch canker and fusiform rust. A total of 23,373 phenotypic data points were collected for lesion length (pitch canker) and gall score, gall length, and gall width (fusiform rust). We verified heritable fusiform rust and pitch canker traits and calculated parental, clonal, and full-sib family rankings for both diseases. Genetic correlations revealed that traits associated with fusiform rust are genetically distinct from one another, and that the genetic mechanisms underlying pitch canker and fusiform rust resistance are independent. The disease phenotyping described here is a critical step towards identifying specific loci and alleles associated with fusiform rust and pitch canker resistance.