Characterization of a new greenbug resistance gene Gb9 in a synthetic hexaploid wheat.

Greenbug [Schizaphis graminum (Rondani)] is a serious insect pest that not only damages cereal crops, but also transmits several destructive viruses. The emergence of new greenbug biotypes in the field makes it urgent to identify novel greenbug resistance genes in wheat. CWI 76364 (PI 703397), a synthetic hexaploid wheat (SHW) line, exhibits greenbug resistance. Evaluation of an F2:3 population from cross OK 14319 × CWI 76364 indicated that a dominant gene, designated Gb9, conditions greenbug resistance in CWI 76364. Selective genotyping of a subset of F2 plants with contrasting phenotypes by genotyping-by-sequencing identified 25 SNPs closely linked to Gb9 on chromosome arm 7DL. Ten of these SNPs were converted to Kompetitive allele-specific polymerase chain reaction (KASP) markers for genotyping the entire F2 population. Genetic analysis delimited Gb9 to a 0.6-Mb interval flanked by KASP markers located at 599,835,668 bp (Stars-KASP872) and 600,471,081 bp (Stars-KASP881) on 7DL. Gb9 was 0.5 cM distal to Stars-KASP872 and 0.5 cM proximal to Stars-KASP881. Allelism tests indicated that Gb9 is a new greenbug resistance gene which confers resistance to greenbug biotypes C, E, H, I, and TX1. TX1 is one of the most widely virulent biotypes and has overcome most known wheat greenbug resistance genes. The introgression of Gb9 into locally adapted wheat cultivars is of economic importance, and the KASP markers developed in this study can be used to tag Gb9 in cultivar development.

Characterization of Quantitative Trait Loci for Leaf Rust Resistance in the Uzbekistani Wheat Landrace Teremai Bugdai.

Leaf rust, caused by Puccinia triticina, is a major cause of wheat yield losses globally, and novel leaf rust resistance genes are needed to enhance wheat leaf rust resistance. Teremai Bugdai is a landrace from Uzebekistan that is highly resistant to many races of P. triticina in the United States. To unravel leaf rust resistance loci in Teremai Bugdai, a recombinant inbred line (RIL) population of Teremai Bugdai × TAM 110 was evaluated for response to P. triticina race Pt54-1 (TNBGJ) and genotyped using single nucleotide polymorphism (SNP) markers generated by genotyping-by-sequencing (GBS). Quantitative trait loci (QTL) analysis using 5,130 high-quality GBS-SNPs revealed three QTLs, QLr-Stars-2DS, QLr-Stars-6BL, and QLr.Stars-7BL, for leaf rust resistance in two experiments. QLr-Stars-2DS, which is either a new Lr2 allele or a new resistance locus, was delimited to an ∼19.47-Mb interval between 46.4 and 65.9 Mb on 2DS and explained 31.3 and 33.2% of the phenotypic variance in the two experiments. QLr-Stars-6BL was mapped in an ∼84.0-kb interval between 719.48 and 719.56 Mb on 6BL, accounting for 33 to 36.8% of the phenotypic variance in two experiments. QLr.Stars-7BL was placed in a 350-kb interval between 762.41 and 762.76 Mb on 7BL and explained 4.4 to 5.3% of the phenotypic variance. Nine GBS-SNPs flanking these QTLs were converted to kompetitive allele specific PCR (KASP) markers, and these markers can be used to facilitate their introgression into locally adapted wheat lines.

Identification of a Novel Pm65 Allele Conferring a Wide Spectrum of Resistance to Powdery Mildew in Wheat Accession PI 351817.

Powdery mildew is caused by the highly adaptive biotrophic fungus Blumeria graminis f. sp. tritici infecting wheat worldwide. Novel powdery mildew resistance genes are urgently needed that can be used rapidly in wheat cultivar development with minimal disruption of trait advances elsewhere. PI 351817 is a German cultivar exhibiting a wide spectrum of resistance to B. graminis f. sp. tritici isolates collected from different wheat-growing regions of the United States. Evaluation of an F2 population and 237 F2:3 lines derived from OK1059060-2C14 × PI 351817 for responses to B. graminis f. sp. tritici isolate OKS(14)-B-3-1 identified a single dominant gene, designated Pm351817, for powdery mildew resistance in PI 351817. Using bulked segregant analysis (BSA) and simple sequence repeat (SSR) markers, Pm351817 was mapped in the terminal region of the long arm of chromosome 2A. Deep sequencing of the genotyping-by-sequencing libraries of the two parental lines identified a set of single-nucleotide polymorphism (SNP) markers in the 2AL candidate gene region. Those SNP markers was subsequently converted to Kompetitive allele-specific PCR (KASP) markers for genotyping the mapping population. Linkage analysis delimited Pm351817 to a 634-kb interval between Stars-KASP656 (771,207,512 bp) and Stars-KASP662 (771,841,609 bp) on 2AL, based on the Chinese Spring reference sequence IWGSC RefSeq v 2.1. Tests of allelism indicated that Pm351817 is located at the Pm65 locus. Pm351817 shows resistance to all B. graminis f. sp. tritici isolates used in this study and can be used to enhance powdery mildew resistance in the United States. KASP markers flanking Pm351817 can be used to select Pm351817 in wheat breeding programs after further tests for polymorphism.

Identification and characterization of the novel leaf rust resistance gene Lr81 in wheat.

KEY MESSAGE: The novel, leaf rust seedling resistance gene, Lr81, was identified in a Croatian breeding line and mapped to a genomic region of less than 100 Kb on chromosome 2AS. Leaf rust, caused by Puccinia triticina, is the most common and widespread rust disease in wheat. Races of Puccinia triticina evolve rapidly in the southern Great Plains of the USA, and leaf rust resistance genes often lose effectiveness shortly after deployment in wheat production. PI 470121, a wheat breeding line developed by the University of Zagreb in Croatia, showed high resistance to Puccinia triticina races collected from Oklahoma, suggesting that PI 470121 could be a leaf rust resistance source for the southern Great Plains of the USA. Genetic analysis based on an F2 population and F2:3 families derived from the cross PI 470121 × Stardust indicated that PI 470121 carries a dominant seedling resistance gene, designated as Lr81. Linkage mapping delimited Lr81 to a genomic region of 96,148 bp flanked by newly developed KASP markers Xstars-KASP320 and Xstars-KASP323 on the short arm of chromosome 2A, spanning 67,030,206-67,132,354 bp in the Chinese Spring reference assembly (IWGSC RefSeq v1.0). Deletion bin mapping assigned Lr81 to the terminal bin 2AS-0.78-1.00. Allelism tests indicated that Lr81 is a distinctive leaf rust resistance locus with the physical order Lr65-Lr17-Lr81. Marker-assisted selection based on a set of markers closely linked to leaf rust resistance genes in PI 470121 and Stardust enabled identification of a recombinant inbred line RIL92 carrying Lr81 only. Lr81 is a valuable leaf rust resistance source that can be rapidly introgressed into locally adapted cultivars using KASP markers Xstars-KASP320 and Xstars-KASP323.

Characterization of an Incomplete Leaf Rust Resistance Gene on Chromosome 1RS and Development of KASP Markers for Lr47 in Wheat.

Leaf rust, caused by Puccinia triticina, is one of the most common wheat (Triticum aestivum) diseases in the Great Plains of the United States. A population of recombinant inbred lines from CI 17884 × 'Bainong 418' was evaluated for responses to leaf rust race Pt52-2 and genotyped using single nucleotide polymorphism (SNP) markers. Quantitative trait locus analysis identified a minor gene for resistance to leaf rust, designated QLr.stars-1RS, on the 1BL.1RS translocation segment in 'Bainong 418', and another leaf rust resistance gene, Lr47, on chromosome 7A of CI 17884. Lr47, originally identified in CI 17884 and located in a wheat-T. speltoides translocation segment 7S#1S, remains one of only a few race-specific resistance genes still effective in the Great Plains. A set of 7A-specific simple sequence repeat markers were developed and used to genotype CI 17884 and a pair of near-isogenic lines differing in the presence or absence of 7S#1S, PI 603918, and 'Pavon F76'. Haplotype analysis indicated that the estimated length of 7S#1S was 157.23 to 174.42 Megabases, accounting for ∼23% of the 7A chromosome. Two SNPs on 7S#1S and four SNPs on the 1RS chromosome arm were converted to Kompetitive allele-specific PCR (KASP) markers, which were subsequently validated in a panel of cultivars and elite breeding lines released within the last decade. Of these, one- and two-KASP markers are specific to the 1RS chromosome arm and 7S#1S, respectively, indicating that they can facilitate the introgression of Lr47 and QLr.stars-1RS into locally adapted wheat cultivars and breeding lines.

Gb8, a new gene conferring resistance to economically important greenbug biotypes in wheat.

KEY MESSAGE: A new greenbug resistance gene Gb8 conferring broad resistance to US greenbug biotypes was identified in hard red winter wheat line PI 595379-1 and was mapped to the terminal region of chromosome 7DL. Greenbug [Schizaphis graminum (Rondani)] is a worldwide insect pest that poses a serious threat to wheat production. New greenbug resistance genes that can be readily used in wheat breeding are urgently needed. The objective of this study was to characterize a greenbug resistance gene in PI 595379-1, a single plant selection from PI 595379. Genetic analysis of response to greenbug biotype E in an F2:3 population derived from a cross between PI 595379-1 and PI 243735 indicated that a single gene, designated Gb8, conditioned resistance. Linkage analysis placed Gb8 in a 2.7-Mb interval in the terminal bin of chromosome 7DL (7DL3-082-1.0), spanning 595.6 to 598.3 Mb in the Chinese Spring IWGSC RefSeq version 1.0 reference sequence. Gb8 co-segregated with a newly developed SSR marker Xstars508, positioned at 596.4 Mb in the reference sequence. Allelism tests showed that Gb8 was different from three permanently named genes on the same chromosome arm and the estimated genetic distance between Gb8 and Gb3 was 15.35 ± 1.35 cM. Gb8 can be directly used in wheat breeding to enhance greenbug resistance.

Development and deployment of KASP markers for multiple alleles of Lr34 in wheat.

KEY MESSAGE: Heterogeneous Lr34 genes for leaf rust in winter wheat cultivar 'Duster' and KASP markers for allelic variation in exon 11 and exon 22 of Lr34. Wheat, Triticum aestivum (2n = 6x = 42, AABBDD), is a hexaploid species, and each of three homoeologous genomes A, B, and D should have one copy for a gene in its ancestral form if the gene has no duplication. Previously reported leaf rust resistance gene Lr34 has one copy on the short arm of chromosome 7D in hexaploid wheat, and allelic variation in Lr34 is in intron 4, exon 11, exon 12, or exon 22. In this study, we discovered that Oklahoma hard red winter wheat cultivar 'Duster' (PI 644,016) has two copies of the Lr34 gene, the resistance allele Lr34a and the susceptibility allele Lr34b. Both Lr34a and Lr34b were mapped in the same linkage group on chromosome 7D in a doubled-haploid population generated from a cross between Duster and a winter wheat cultivar 'Billings' which carries the susceptibility allele Lr34c. A chromosomal fragment including Lr34 and at least two neighboring genes on its proximal side but excluding genes on its distal side was duplicated in Duster. The Duster Lr34ab allele was associated with tip necrosis and increased resistance against leaf rust at adult plants in the Duster × Billings DH population tested in the field, demonstrating the function of the Duster Lr34ab allele in wheat. We have developed KASP markers for allelic variation in exon 11 and exon 22 of Lr34 in wheat. These markers can be utilized to accelerate the selection of Lr34 in wheat.

Characterization of Pm63, a powdery mildew resistance gene in Iranian landrace PI 628024.

KEY MESSAGE: A new powdery mildew resistance gene conferring a wide spectrum of resistance to Bgt isolates in the USA, Pm63 , was identified in Iranian wheat landrace PI 628024 and mapped to the terminal region of the long arm of chromosome 2B. Powdery mildew is a globally important wheat disease causing severe yield losses, and host resistance is the preferred strategy for managing this disease. The objective of this study was to characterize a powdery mildew resistance gene in Iranian landrace PI 628024, which exhibited a wide spectrum of resistance to representative Blumeria graminis f. sp. tritici (Bgt) isolates collected from different regions of the USA. An F2 population and F2:3 lines derived from the cross PI 628024 × CItr 11349 were used in this study, and genetic analysis indicated that a single dominant gene, designated Pm63, conferred resistance to Bgt isolate OKS(14)-B-3-1. Linkage analysis located Pm63 to an interval of about 13.1 Mb on the long arm of chromosome 2B, spanning 710.3-723.4 Mb in the Chinese Spring reference sequence. Bin mapping assigned Pm63 to the terminal bin 2BL6-0.89-1.0, 1.1 cM proximal to STS marker Xbcd135-2 and 0.6 cM distal to SSR marker Xstars419. Allelism tests indicated that Pm63 is a new powdery mildew resistance gene, which differs from other genes in the terminal bin by origin, genomic location, and responses to a set of 16 representative US Bgt isolates. Pm63 can be widely used to enhance powdery mildew resistance in the Great Plains, western, and southeastern regions of the USA.

Characterization of Pm65, a new powdery mildew resistance gene on chromosome 2AL of a facultative wheat cultivar.

KEY MESSAGE: A new powdery mildew resistance gene that can be readily used in wheat breeding, Pm65, was identified in the facultative wheat cultivar Xinmai 208 and mapped to the terminal region of chromosome 2AL. Wheat powdery mildew, a widely occurring disease caused by the biotrophic fungus Blumeriagraminis f. sp. tritici (Bgt), poses a serious threat to wheat production. A high breeding priority is to identify powdery mildew resistance genes that can be readily used either alone or in gene complexes involving other disease resistance genes. An F2 population and 227 F2:3 families derived from the cross Xinmai 208 × Stardust were generated to map a powdery mildew resistance gene in Xinmai 208, a high-yielding Chinese wheat cultivar. Genetic analysis indicated that Xinmai 208 carries a single dominant powdery mildew resistance gene, designated herein Pm65, and linkage analysis delimited Pm65 to a 0.5 cM interval covering 531.8 Kb (763,289,667-763,821,463 bp) on chromosome 2AL in the Chinese Spring reference sequence. An allelism test indicated that Pm65 is a new gene about 10.3 cM distal to the Pm4 locus. Pm65 was 0.3 cM proximal to Xstars355 and 0.2 cM distal to Xstars356. It conferred near-immunity to 19 of 20 Bgt isolates collected from different wheat-growing regions of the USA. Coming from a high-yield potential cultivar, Pm65 can be directly used to enhance powdery mildew resistance in wheat. The newly developed SSR markers Xstars355 and Xstars356 have the potential to tag Pm65 for wheat improvement.

Nitrogen use efficiency is regulated by interacting proteins relevant to development in wheat.

Wheat (Triticum aestivum) has low nitrogen use efficiency (NUE). The genetic mechanisms controlling NUE are unknown. Positional cloning of a major quantitative trait locus for N-related agronomic traits showed that the vernalization gene TaVRN-A1 was tightly linked with TaNUE1, the gene shown to influence NUE in wheat. Because of an Ala180 /Val180 substitution, TaVRN-A1a and TaVRN-A1b proteins interact differentially with TaANR1, a protein encoded by a wheat orthologue of Arabidopsis nitrate regulated 1 (ANR1). The transcripts of both TaVRN-A1 and TaANR1 were down-regulated by nitrogen. TaANR1 was functionally characterized in TaANR1::RNAi transgenic wheat, and in a natural mutant with a 23-bp deletion including 10-bp at the 5' end of intron 5 and 13-bp of exon 6 in gDNA sequence in its gDNA sequence, which produced transcript that lacked the full 84-bp exon 6. Both TaANR1 and TaHOX1 bound to the Ala180 /Val180 position of TaVRN-A1. Genetically incorporating favourable alleles from TaVRN-A1, TaANR1 and TaHOX1 increased grain yield from 9.84% to 11.58% in the field. Molecular markers for allelic variation of the genes that regulate nitrogen can be used in breeding programmes aimed at improving NUE and yield in novel wheat cultivars.

Pm223899, a new recessive powdery mildew resistance gene identified in Afghanistan landrace PI 223899.

KEY MESSAGE: A new recessive powdery mildew resistance gene, Pm223899, was identified in Afghanistan wheat landrace PI 223899 and mapped to an interval of about 831 Kb in the terminal region of the short arm of chromosome 1A. Wheat powdery mildew, a globally important disease caused by the biotrophic fungus Blumeria graminis f.sp. tritici (Bgt), has occurred with increased frequency and severity in recent years, and some widely deployed resistance genes have lost effectiveness. PI 223899 is an Afghanistan landrace exhibiting high resistance to Bgt isolates collected from the Great Plains. An F2 population and F2:3 lines derived from a cross between PI 223899 and OK1059060-126135-3 were evaluated for response to Bgt isolate OKS(14)-B-3-1, and the bulked segregant analysis (BSA) approach was used to map the powdery mildew resistance gene. Genetic analysis indicated that a recessive gene, designated Pm223899, conferred powdery mildew resistance in PI 223899. Linkage analysis placed Pm223899 to an interval of about 831 Kb in the terminal region of chromosome 1AS, spanning 4,504,697-5,336,062 bp of the Chinese Spring reference sequence. Eight genes were predicted in this genomic region, including TraesCS1AG008300 encoding a putative disease resistance protein RGA4. Pm223899 was flanked proximally by a SSR marker STARS333 (1.4 cM) and distally by the Pm3 locus (0.3 cM). One F2 recombinant was identified between Pm3 and Pm223899 using a Pm3b-specific marker, indicating that Pm223899 is most likely a new gene, rather than an allele of the Pm3 locus. Pm223389 confers a high level of resistance to Bgt isolates collected from Pennsylvania, Oklahoma, Nebraska, and Montana. Therefore, Pm223389 can be used to enhance powdery mildew resistance in these states. Pm3b-1 and STARS333 have the potential to tag Pm223389 in wheat breeding.

Correction to: Pm223899, a new recessive powdery mildew resistance gene identified in Afghanistan landrace PI 223899.

Unfortunately, the caption of Fig. 2 was incorrectly published in the original publication. The complete correct caption should read as follows.

Characterization of Pm59, a novel powdery mildew resistance gene in Afghanistan wheat landrace PI 181356.

KEY MESSAGE: A new powdery mildew resistance gene, designated Pm59, was identified in Afghanistan wheat landrace PI 181356, and mapped in the terminal region of the long arm of chromosome 7A. Powdery mildew, caused by Blumeria graminis f. sp. tritici (Bgt), is an important foliar disease of wheat worldwide. In the Great Plains of the USA, Bgt isolates virulent to widely used powdery mildew resistance genes, such as Pm3a, were previously identified. The objectives of this study were to characterize the powdery mildew resistance gene in Afghanistan landrace PI 181356, which exhibited high resistance to Bgt isolates collected in southern Great Plains, and identify molecular markers for marker-assisted selection. An F2 population and F2:3 lines derived from a cross between PI 181356 and OK1059060-126135-3 were used in this study. Genetic analysis indicated that PI 181356 carries a single dominant gene, designated Pm59, in the terminal region of the long arm of chromosome 7A. Pm59 was mapped to an interval between sequence tag site (STS) markers Xmag1759 and Xmag1714 with genetic distances of 0.4 cM distal to Xmag1759 and 5.7 cM proximal to Xmag1714. Physical mapping suggested that Pm59 is in the distal bin 7AL 0.99-1.00. Pm59 is a novel powdery mildew resistance gene, and confers resistance to Bgt isolates collected from the Great Plains and the state of Montana. Therefore, Pm59 can be used to breed powdery mildew-resistant cultivars in these regions. Xmag1759 is ideal for marker-assisted selection of Pm59 in wheat breeding.

Genetic basis of the very short life cycle of 'Apogee' wheat.

BACKGROUND: 'Apogee' has a very short life cycle among wheat cultivars (flowering 25 days after planting under a long day and without vernalization), and it is a unique genetic material that can be used to accelerate cycling breeding lines. However, little is known about the genetic basis of the super-short life of Apogee wheat. RESULTS: In this study, Apogee was crossed with a strong winter wheat cultivar 'Overland', and 858 F2 plants were generated and tested in a greenhouse under constant warm temperature and long days. Apogee wheat was found to have the early alleles for four flowering time genes, which were ranked in the order of vrn-A1 > VRN-B1 > vrn-D3 > PPD-D1 according to their effect intensity. All these Apogee alleles for early flowering showed complete or partial dominance effects in the F2 population. Surprisingly, Apogee was found to have the same alleles at vrn-A1a and vrn-D3a for early flowering as observed in winter wheat cultivar 'Jagger.' It was also found that the vrn-A1a gene was epistatic to VRN-B1 and vrn-D3. The dominant vrn-D3a alone was not sufficient to cause the transition from vegetative to reproductive development in winter plants without vernalization but was able to accelerate flowering in those plants that carry the vrn-A1a or Vrn-B1 alleles. The genetic effects of the vernalization and photoperiod genes were validated in Apogee x Overland F3 populations. CONCLUSION: VRN-A1, VRN-B1, VRN-D3, and PPD-D1 are the major genes that enabled Apogee to produce the very short life cycle. This study greatly advanced the molecular understanding of the multiple flowering genes under different genetic backgrounds and provided useful molecular tools that can be used to accelerate winter wheat breeding schemes.

Genome-wide association study reveals genetic architecture of coleoptile length in wheat.

KEY MESSAGE: Eight QTL for coleoptile length were identified in a genome-wide association study on a set of 893 wheat accessions, four of which are novel loci. Wheat cultivars with long coleoptiles are preferred in wheat-growing regions where deep planting is practiced. However, the wide use of gibberellic acid (GA)-insensitive dwarfing genes, Rht-B1b and Rht-D1b, makes it challenging to breed dwarf wheat cultivars with long coleoptiles. To understand the genetic basis of coleoptile length, we performed a genome-wide association study on a set of 893 landraces and historical cultivars using 5011 single nucleotide polymorphism (SNP) markers. Structure analysis revealed four subgroups in the association panel. Association analysis results suggested that Rht-B1b and Rht-D1b genes significantly reduced coleoptile length, and eight additional quantitative trait loci (QTL) for coleoptile length were also identified. These QTL explained 1.45-3.18 and 1.36-3.11% of the phenotypic variation in 2015 and 2016, respectively, and their allelic substitution effects ranged from 0.31 to 1.75 cm in 2015, and 0.63-1.55 cm in 2016. Of the eight QTL, QCL.stars-1BS1, QCL.stars-2DS1, QCL.stars-4BS2, and QCL.stars-5BL1 are likely novel loci for coleoptile length. The favorable alleles in each accession ranged from two to eight with an average of 5.8 at eight loci in the panel, and more favorable alleles were significantly associated with longer coleoptile, suggesting that QTL pyramiding is an effective approach to increase wheat coleoptile length.

Precisely mapping a major gene conferring resistance to Hessian fly in bread wheat using genotyping-by-sequencing.

BACKGROUND: One of the reasons hard red winter wheat cultivar 'Duster' (PI 644016) is widely grown in the southern Great Plains is that it confers a consistently high level of resistance to biotype GP of Hessian fly (Hf). However, little is known about the genetic mechanism underlying Hf resistance in Duster. This study aimed to unravel complex structures of the Hf region on chromosome 1AS in wheat by using genotyping-by-sequencing (GBS) markers and single nucleotide polymorphism (SNP) markers. RESULTS: Doubled haploid (DH) lines generated from a cross between two winter wheat cultivars, 'Duster' and 'Billings' , were used to identify genes in Duster responsible for effective and consistent resistance to Hf. Segregation in reaction of the 282 DH lines to Hf biotype GP fit a one-gene model. The DH population was genotyped using 2,358 markers developed using the GBS approach. A major QTL, explaining 88% of the total phenotypic variation, was mapped to a chromosome region that spanned 178 cM and contained 205 GBS markers plus 1 SSR marker and 1 gene marker, with 0.86 cM per marker in genetic distance. The analyses of GBS marker sequences and further mapping of SSR and gene markers enabled location of the QTL-containing linkage group on the short arm of chromosome 1A. Comparative mapping of the common markers for the gene for QHf.osu-1A (d) in Duster and the Hf-resistance gene for QHf.osu-1A (74) in cultivar '2174' showed that the two Hf resistance genes are located on the same chromosome arm 1AS, only 11.2 cM apart in genetic distance. The gene at QHf.osu-1A (d) in Duster has been delimited within a 2.7 cM region. CONCLUSION: Two distinct resistance genes exist on the short arm of chromosome 1A as found in the two hard red winter cultivars, 2174 and Duster. Whereas the Hf resistance gene in 2174 is likely allelic to one or more of the previously mapped resistance genes (H9, H10, H11, H16, or H17) in wheat, the gene in Duster is novel and confers a more consistent phenotype than 2174 in response to biotype GP infestation in controlled-environment assays.

Vernalization requirement duration in winter wheat is controlled by TaVRN-A1 at the protein level.

Winter wheat requires a period of low temperatures to accelerate flowering (vernalization). This requirement could make winter wheat more vulnerable to elevated global temperature via insufficient vernalization. All known vernalization genes are cloned according to qualitative variation in vernalization requirement between spring and winter wheat, but the genes controlling quantitative variation for more or less vernalization requirement among winter wheat cultivars remain unknown. We report here that the gene for the vernalization requirement duration in winter wheat was cloned using a BC(1)F(2:3) population that segregated in a 3:1 ratio of early-flowering plants and late-flowering plants after vernalization for 3 weeks. The positional cloning of the gene for vernalization requirement duration demonstrated that this trait is controlled by TaVRN-A1 at the protein level. The Ala(180) in vrn-A1a, encoded by the dominant allele for 3-week vernalization, was mutated to Val(180) in vrn-A1b, encoded by the recessive allele for 6-week vernalization. Further studies with in vitro protein pull-down assays and immunoprecipitation analyses indicated that the mutated Val(180) in vrn-A1b protein decreased the ability to bind with TaHOX1 (the first homeobox protein in Triticum aestivum). The direct binding of TaVRN-A1 and TaHOX1 proteins was confirmed in the nucleus of living plant cells by bimolecular fluorescence complementation (BiFC) analyses. The TaHOX1 gene was found to be upregulated by low temperatures, and to have a significant genetic effect on heading date, suggesting that TaHOX1 functions in the flowering pathway in winter wheat.

Identification of a new Rsg1 allele conferring resistance to multiple greenbug biotypes from barley accessions PI 499276 and PI 566459.

Greenbug [Schizaphis graminum (Rondani)] is a major insect pest that significantly affects barley production worldwide. The identification of novel greenbug resistance genes is crucial for sustainable barley production and global food security. To identify greenbug resistance genes from a US breeding line PI 499276 and a Chinese cultivar PI 566459, two F6:7 recombinant inbred line (RIL) populations developed from crosses Weskan × PI 499276 and Weskan × PI 566459 were phenotyped for responses to greenbug biotype E and genotyped using genotyping-by-sequencing (GBS). Linkage analysis using single nucleotide polymorphism and kompetitive allele-specific polymorphism (KASP) markers delimited the greenbug resistance genes from PI 499276 and PI 566459 to a 1.2 Mb genomic region between 666.5 and 667.7 Mb on the long arm of chromosome 3H in the Morex Hordeum vulgare r1 reference sequence. Allelism tests based on responses of four F2 populations to greenbug biotype E indicated that the greenbug resistance gene in PI 499276 and PI 566459 is either allelic or very close to Rsg1. Given that PI 499276 and PI 566459 shared the same unique resistance pattern to a set of 14 greenbug biotypes, which is different from those of other Rsg1 alleles, they carry a new Rsg1 allele. The greenbug resistance genes in Post 90, PI 499276/PI 566459, and WBDC 336 were designated as Rsg1.a1, Rsg1.a2, and Rsg1.a3, respectively. KASP markers KASP-Rsg1a3-1, KASP-Rsg1a3-2, and KASP160 can be used to tag Rsg1.a2 in barley breeding.

Characterization of Rsg3, a novel greenbug resistance gene from the Chinese barley landrace PI 565676.

Greenbug (Schizaphis graminum Rondani) is a pest that poses a serious threat to cereal production worldwide. Yield losses caused by greenbug are predicted to increase because of global warming. To date, only a few barley (Hordeum vulgare L.) greenbug resistance genes have been reported and new genes are urgently needed because of the continuous occurrence of novel greenbug biotypes. PI 565676, a landrace collected from Henan province of China, exhibits high resistance to several predominant greenbug biotypes. An F6:7 recombinant inbred line (RIL) population derived from the cross PI 565676 × 'Weskan' was evaluated for response to greenbug biotypes E and F using a standard aphid assay protocol, and a randomized complete block design with two replicates was adopted. The RIL population was genotyped using single-nucleotide polymorphisms (SNPs) markers generated by genotyping-by-sequencing (GBS). Gene mapping placed the greenbug resistance gene in PI 565676, designated Rsg3, to an interval of 93,140 bp between 667,558,306 and 667,651,446 bp on the long arm of chromosome 3H. Four high-confidence genes were annotated in this region with one encoding a leucine-rich repeat-containing protein. An allelism test indicated that Rsg3 is independent of the Rsg1 locus, with estimated recombination frequency of 12.85 ± 0.20% and genetic distance of 13.14 ± 0.21 cM between the two loci. Therefore, Rsg3 represents a new locus for greenbug resistance. Two SNPs flanking Rsg3 were converted to Kompetitive Allele Specific PCR (KASP) markers, which can be used to tag Rsg3 in barley breeding.

O-linked N-acetylglucosamine transferase is involved in fine regulation of flowering time in winter wheat.

Vernalization genes underlying dramatic differences in flowering time between spring wheat and winter wheat have been studied extensively, but little is known about genes that regulate subtler differences in flowering time among winter wheat cultivars, which account for approximately 75% of wheat grown worldwide. Here, we identify a gene encoding an O-linked N-acetylglucosamine (O-GlcNAc) transferase (OGT) that differentiates heading date between winter wheat cultivars Duster and Billings. We clone this TaOGT1 gene from a quantitative trait locus (QTL) for heading date in a mapping population derived from these two bread wheat cultivars and analyzed in various environments. Transgenic complementation analysis shows that constitutive overexpression of TaOGT1b from Billings accelerates the heading of transgenic Duster plants. TaOGT1 is able to transfer an O-GlcNAc group to wheat protein TaGRP2. Our findings establish important roles for TaOGT1 in winter wheat in adaptation to global warming in the future climate scenarios.