Transcriptome and metabolome analyses reveal molecular insights into waterlogging tolerance in Barley.

Waterlogging stress is one of the major abiotic stresses affecting the productivity and quality of many crops worldwide. However, the mechanisms of waterlogging tolerance are still elusive in barley. In this study, we identify key differentially expressed genes (DEGs) and differential metabolites (DM) that mediate distinct waterlogging tolerance strategies in leaf and root of two barley varieties with contrasting waterlogging tolerance under different waterlogging treatments. Transcriptome profiling revealed that the response of roots was more distinct than that of leaves in both varieties, in which the number of downregulated genes in roots was 7.41-fold higher than that in leaves of waterlogging sensitive variety after 72 h of waterlogging stress. We also found the number of waterlogging stress-induced upregulated DEGs in the waterlogging tolerant variety was higher than that of the waterlogging sensitive variety in both leaves and roots in 1 h and 72 h treatment. This suggested the waterlogging tolerant variety may respond more quickly to waterlogging stress. Meanwhile, phenylpropanoid biosynthesis pathway was identified to play critical roles in waterlogging tolerant variety by improving cell wall biogenesis and peroxidase activity through DEGs such as Peroxidase (PERs) and Cinnamoyl-CoA reductases (CCRs) to improve resistance to waterlogging. Based on metabolomic and transcriptomic analysis, we found the waterlogging tolerant variety can better alleviate the energy deficiency via higher sugar content, reduced lactate accumulation, and improved ethanol fermentation activity compared to the waterlogging sensitive variety. In summary, our results provide waterlogging tolerance strategies in barley to guide the development of elite genetic resources towards waterlogging-tolerant crop varieties.

Genome-wide association studies reveal stable loci for wheat grain size under different sowing dates.

Background: Wheat (Tritium aestivum L.) production is critical for global food security. In recent years, due to climate change and the prolonged growing period of rice varieties, the delayed sowing of wheat has resulted in a loss of grain yield in the area of the middle and lower reaches of the Yangtze River. It is of great significance to screen for natural germplasm resources of wheat that are resistant to late sowing and to explore genetic loci that stably control grain size and yield. Methods: A collection of 327 wheat accessions from diverse sources were subjected to genome-wide association studies using genotyping-by-sequencing. Field trials were conducted under normal, delayed, and seriously delayed sowing conditions for grain length, width, and thousand-grain weight at two sites. Additionally, the additive main effects and multiplicative interaction (AMMI) model was applied to evaluate the stability of thousand-grain weight of 327 accessions across multiple sowing dates. Results: Four wheat germplasm resources have been screened, demonstrating higher stability of thousand-grain weight. A total of 43, 35, and 39 significant MTAs were determined across all chromosomes except for 4D under the three sowing dates, respectively. A total of 10.31% of MTAs that stably affect wheat grain size could be repeatedly identified in at least two sowing dates, with PVE ranging from 0.03% to 38.06%. Among these, six were for GL, three for GW, and one for TGW. There were three novel and stable loci (4A_598189950, 4B_307707920, 2D_622241054) located in conserved regions of the genome, which provide excellent genetic resources for pyramid breeding strategies of superior loci. Our findings offer a theoretical basis for cultivar improvement and marker-assisted selection in wheat breeding practices.

Genome-wide association studies reveal novel loci for grain size in two-rowed barley (Hordeum vulgare L.).

KEY MESSAGE: SNP-based and InDel-based GWAS on multi-environment data identified genomic regions associated with barley grain size. Barley yield and quality are greatly influenced by grain size. Improving barley grain size in breeding programs requires knowledge of genetic loci and alleles in germplasm resources. In this study, a collection of 334 worldwide two-rowed barley accessions with extensive genetic diversity was evaluated for grain size including grain length (GL), grain width (GW), and thousand-grain weight (TGW) across six independent field trials. Significant differences were observed in genotype and environments for all measured traits. SNP- and InDel-based GWAS were applied to dissect the genetic architecture of grain size with an SLAF-seq strategy. Two approaches using the FarmCPU model revealed 38 significant marker-trait associations (MTAs) with PVE ranging from 0.01% to 20.68%. Among these MTAs, five were on genomic regions where no previously reported QTL for grain size. Superior alleles of TGW-associated SNP233060 and GL-associated InDel11006 exhibited significantly higher levels of phenotype. The significant MTAs could be used in marker-assisted selection breeding.

Transcriptomic and Metabolomic Analyses Reveal the Importance of Lipid Metabolism and Photosynthesis Regulation in High Salinity Tolerance in Barley (Hordeum vulgare L.) Leaves Derived from Mutagenesis Combined with Microspore Culture.

Barley is the most salt-tolerant cereal crop. However, little attention has been paid to the salt-tolerant doubled haploids of barley derived from mutagenesis combined with isolated microspore culture. In the present study, barley doubled haploid (DH) line 20, which was produced by mutagenesis combined with isolated microspore culture, showed stably and heritably better salt tolerance than the wild type H30 in terms of fresh shoot weight, dry shoot weight, K+/Na+ ratio and photosynthetic characteristics. Transcriptome and metabolome analyses were performed to compare the changes in gene expression and metabolites between DH20 and H30. A total of 462 differentially expressed genes (DEGs) and 152 differentially accumulated metabolites (DAMs) were identified in DH20 compared to H30 under salt stress. Among the DAMs, fatty acids were the most accumulated in DH20 under salt stress. The integration of transcriptome and metabolome analyses revealed that nine key biomarkers, including two metabolites and seven genes, could distinguish DH20 and H30 when exposed to high salt. The pathways of linoleic acid metabolism, alpha-linolenic acid metabolism, glycerolipid metabolism, photosynthesis, and alanine, aspartate and glutamate metabolism were significantly enriched in DH20 with DEGs and DAMs in response to salt stress. These results suggest that DH20 may enhance resilience by promoting lipid metabolism, maintaining energy metabolism and decreasing amino acids metabolism. The study provided novel insights for the rapid generation of homozygous mutant plants by mutagenesis combined with microspore culture technology and also identified candidate genes and metabolites that may enable the mutant plants to cope with salt stress.

Genome-wide analysis of the MADS-box gene family involved in salt and waterlogging tolerance in barley (Hordeum vulgare L.).

MADS-box transcription factors are crucial members of regulatory networks underlying multiple developmental pathways and abiotic stress regulatory networks in plants. Studies on stress resistance-related functions of MADS-box genes are very limited in barley. To gain insight into this gene family and elucidate their roles in salt and waterlogging stress resistance, we performed genome-wide identification, characterization and expression analysis of MADS-box genes in barley. A whole-genome survey of barley revealed 83 MADS-box genes, which were categorized into type I (Mα, Mß and Mγ) and type II (AP1, SEP1, AGL12, STK, AGL16, SVP and MIKC*) lineages based on phylogeny, protein motif structure. Twenty conserved motifs were determined and each HvMADS contained one to six motifs. We also found tandem repeat duplication was the driven force for HvMADS gene family expansion. Additionally, the co-expression regulatory network of 10 and 14 HvMADS genes was predicted in response to salt and waterlogging stress, and we proposed HvMADS11,13 and 35 as candidate genes for further exploration of the functions in abiotic stress. The extensive annotations and transcriptome profiling reported in this study ultimately provides the basis for MADS functional characterization in genetic engineering of barley and other gramineous crops.

Molecular insights into the responses of barley to yellow mosaic disease through transcriptome analysis.

BACKGROUND: Barley (Hordeum vulgare L.) represents the fourth most essential cereal crop in the world, vulnerable to barley yellow mosaic virus (BaYMV) and/or barley mild mosaic virus (BaMMV), leading to the significant yield reduction. To gain a better understanding of the mechanisms regarding barley crop tolerance to virus infection, we employed a transcriptome sequencing approach and investigated global gene expression among three barley varieties under both infected and control conditions. RESULTS: High-throughput sequencing outputs revealed massive genetic responses, reflected by the barley transcriptome after BaYMV and/or BaMMV infection. Significant enrichments in peptidase complex and protein processing in endoplasmic reticulum were clustered through Gene ontology and KEGG analysis. Many genes were identified as transcription factors, antioxidants, disease resistance genes and plant hormones and differentially expressed between infected and uninfected barley varieties. Importantly, general response genes, variety-specific and infection-specific genes were also discovered. Our results provide useful information for future barley breeding to resist BaYMV and BaMMV. CONCLUSIONS: Our study elucidates transcriptomic adaptations in barley response to BaYMV/BaMMV infection through high-throughput sequencing technique. The analysis outcome from GO and KEGG pathways suggests that BaYMV disease induced regulations in multiple molecular-biology processes and signalling pathways. Moreover, critical DEGs involved in defence and stress tolerance mechanisms were displayed. Further functional investigations focusing on these DEGs contributes to understanding the molecular mechanisms of plant response to BaYMV disease infection, thereby offering precious genetic resources for breeding barley varieties resistant to BaYMV disease.

Genetic Localization and Homologous Genes Mining for Barley Grain Size.

Grain size is an important agronomic trait determining barley yield and quality. An increasing number of QTLs (quantitative trait loci) for grain size have been reported due to the improvement in genome sequencing and mapping. Elucidating the molecular mechanisms underpinning barley grain size is vital for producing elite cultivars and accelerating breeding processes. In this review, we summarize the achievements in the molecular mapping of barley grain size over the past two decades, highlighting the results of QTL linkage analysis and genome-wide association studies. We discuss the QTL hotspots and predict candidate genes in detail. Moreover, reported homologs that determine the seed size clustered into several signaling pathways in model plants are also listed, providing the theoretical basis for mining genetic resources and regulatory networks of barley grain size.

HvNCX, a prime candidate gene for the novel qualitative locus qS7.1 associated with salinity tolerance in barley.

KEY MESSAGE: A major QTL (qS7.1) for salinity damage score and Na+ exclusion was identified on chromosome 7H from a barley population derived from a cross between a cultivated variety and a wild accession. qS7.1 was fine-mapped to a 2.46 Mb physical interval and HvNCX encoding a sodium/calcium exchanger is most likely the candidate gene. Soil salinity is one of the major abiotic stresses affecting crop yield. Developing salinity-tolerant varieties is critical for minimizing economic penalties caused by salinity and providing solutions for global food security. Many genes/QTL for salt tolerance have been reported in barley, but only a few of them have been cloned. In this study, a total of 163 doubled haploid lines from a cross between a cultivated barley variety Franklin and a wild barley accession TAM407227 were used to map QTL for salinity tolerance. Four significant QTL were identified for salinity damage scores. One (qS2.1) was located on 2H, determining 7.5% of the phenotypic variation. Two (qS5.1 and qS5.2) were located on 5H, determining 5.3-11.7% of the phenotypic variation. The most significant QTL was found on 7H, explaining 27.8% of the phenotypic variation. Two QTL for Na+ content in leaves under salinity stress were detected on chromosomes 1H (qNa1.1) and 7H(qNa7.1). qS7.1 was fine-mapped to a 2.46 Mb physical interval using F4 recombinant inbred lines. This region contains 23 high-confidence genes, with HvNCX which encodes a sodium/calcium exchanger being most likely the candidate gene. HvNCX was highly induced by salinity stress and showed a greater expression level in the sensitive parent. Multiple nucleotide substitutions and deletions/insertions in the promoter sequence of HvNCX were found between the two parents. cDNA sequencing of the HvNCX revealed that the difference between the two parents is conferred by a single Ala77/Pro77 amino acid substitution, which is located on the transmembrane domain. These findings open new prospects for improving salinity tolerance in barley by targeting a previously unexplored trait.

In situ mapping of ion distribution profiles and gene expression reveals interactions between hypoxia and Mn2+/Fe2+ availability in barley roots.

Flooding stress affects soil properties thus altering the availability, uptake, and transport of mineral nutrients in plant roots. Flooding stress also increases the amount of soluble Mn2+ and Fe2+ in the soil and their uptake by plants, causing elemental toxicity. However, as oxygen profiles in plant roots are not uniform, it is still unclear how soil flooding will affect Mn2+/Fe2+ absorption and distribution in different cell types and tissues. In this study, waterlogging sensitive barley variety NasoNijo (NN) and tolerant variety TX9425 (TX) were exposed to hypoxia, metal (Mn2+ and Fe2+), and combined hypoxia + metal treatment to map the in situ ion profiles at different regions of barley root. We found that combined hypoxia and metal stress causes significantly more reduction in plant biomass compared with the single submergence or metal stress. Despite this, more Fe and Mn were accumulated under metal stress condition than those under combined stress, regardless of variety. Cultivar NN absorbed more Fe and Mn than TX in the cortical cells of the root meristem and in the mature zone under metal stress which was also verified by histochemical detection. In the mature zone, the expressions of Fe and Mn transporter genes including HvADPRibase-Mn (Manganese-dependent ADP-ribose), HvZIP1 (zinc-regulated transporter /Fe-regulated transporter-like protein 1), HvYS1 (yellow stripe 1), HvNRAMP5 (Natural Resistance-Associated Macrophage Protein 5) were significantly downregulated under all three treatments in both barley varieties except HvADPRibase-Mn HvZIP1 cortex of TX were unchanged under metal stress. Interestingly, the transcripts of HvMTP1 (metal tolerance protein 1) were significantly downregulated by metal and combined stress in stele and upregulated by hypoxia and metal stress in cortex of TX, but not affected in NN. It is concluded that Fe and Mn absorption involving HvMTP1is associated with the extent of waterlogging tolerance in barley.

Brittle culm 3, encoding a cellulose synthase subunit 5, is required for cell wall biosynthesis in barley (Hordeum vulgare L.).

The cell wall plays an important role in plant mechanical strength. Cellulose is the major component of plant cell walls and provides the most abundant renewable biomass resource for biofuels on earth. Mutational analysis showed that cellulose synthase (CESA) genes are critical in cell wall biosynthesis in cereal crops like rice. However, their role has not been fully elucidated in barley. In this study, we isolated a brittle culm mutant brittle culm 3 (bc3) derived from Yangnongpi 5 ethyl methanesulfonate (EMS) mutagenesis in barley. The bc3 mutants exhibited reduced mechanical strength of the culms due to impaired thickening of the sclerenchyma cell wall and reduced cellulose and hemicellulose content in the culms. Genetic analysis and map-based cloning revealed that the bc3 mutant was controlled by a single recessive gene and harbored a point mutation in the HvCESA5 gene, generating a premature stop codon near the N-terminal of the protein. Quantitative real-time PCR (qRT-PCR) analysis showed that the HvCESA5 gene is predominantly expressed in the culms and co-expressed with HvCESA4 and HvCESA8, consistent with the brittle culm phenotype of the bc3 mutant. These results indicate that the truncated HvCESA5 affects cell wall biosynthesis leading to a brittle culm phenotype. Our findings provide evidence for the important role of HvCESA5 in cell wall biosynthesis pathway and could be a potential target to modify cell wall in barley.

Mutation of barley HvPDIL5-1 improves resistance to yellow mosaic virus disease without growth or yield penalties.

The soil-borne yellow mosaic virus disease, which is caused by the bymoviruses barley yellow mosaic virus (BaYMV) and/or barley mild mosaic virus (BaMMV), seriously threatens winter barley production in Europe and East Asia. Both viruses are transmitted by the soil-borne plasmodiophorid Polymyxa graminis and are difficult to eliminate through chemical or physical measures in the field, making breeding for resistant cultivars the optimal strategy for disease control. The resistance locus rym1/11 was cloned encoding the host factor gene Protein Disulfide Isomerase Like 5-1 (PDIL5-1), whose loss-of-function variants confer broad-spectrum resistance to multiple strains of BaMMV/BaYMV. Most resistance-conferring variants have been identified in six-rowed barley landraces/historic cultivars, and their introgression into modern two-rowed malting cultivars is difficult because PDIL5-1 is located in a peri-centromeric region with suppressed recombination. In this study, we used CRISPR/Cas9 genome editing to modify PDIL5-1 in the BaYMV/BaMMV-susceptible elite malting barley cv. 'Golden Promise' and obtained the mutants pdil5-1-a and pdil5-1-b. PDIL5-1 in the pdil5-1-a mutant encodes a protein lacking a cysteine residue, and pdil5-1-b contains a protein-coding frameshift. Both mutants were completely resistant to BaYMV. The knockout mutant pdil5-1-b showed complete BaMMV resistance, while pdil5-1-a showed decreased viral accumulation but no disease symptoms if compared to 'Golden Promise'. Both PDIL5-1 edited lines, as well as the previously produced EMS-induced pdil5-1 mutant '10253-1-5' in the elite malting barley cv. 'Barke' background, displayed no growth or yield penalties in garden experiments or bymovirus-free field trials. Line '10253-1-5' showed improved resistance and yield performance compared to the wild-type and its sibling line when grown in infectious fields. Therefore, genome editing of the host factor gene PDIL5-1 could facilitate the breeding of barley varieties with resistance to bymoviruses.

Screening of stable resistant accessions and identification of resistance loci to Barley yellow mosaic virus disease.

Background: The disease caused by Barley yellow mosaic virus (BaYMV) infection is a serious threat to autumn-sown barley (Hordeum vulgare L.) production in Europe, East Asia and Iran. Due to the rapid diversification of BaYMV strains, it is urgent to discover novel germplasm and genes to assist breeding new varieties with resistance to different BaYMV strains, thus minimizing the effect of BaYMV disease on barley cropping. Methods: A natural population consisting of 181 barley accessions with different levels of resistance to BaYMV disease was selected for field resistance identification in two separate locations (Yangzhou and Yancheng, Jiangsu Province, China). Additive main effects and multiplicative interaction (AMMI) analysis was used to identify accessions with stable resistance. Genome-wide association study (GWAS) of BaYMV disease resistance was broadly performed by combining both single nucleotide polymorphisms (SNPs) and specific molecular markers associated with the reported BaYMV disease resistance genes. Furthermore, the viral protein genome linked (VPg) sequences of the virus were amplified and analyzed to assess the differences between the BaYMV strains sourced from the different experimental sites. Results: Seven barley accessions with lower standardized Area Under the Disease Progress Steps (sAUDPS) index in every environment were identified and shown to have stable resistance to BaYMV disease in each assessed location. Apart from the reported BaYMV disease resistance genes rym4 and rym5, one novel resistance locus explaining 24.21% of the phenotypic variation was identified at the Yangzhou testing site, while two other novel resistance loci that contributed 19.23% and 19.79% of the phenotypic variation were identified at the Yancheng testing site, respectively. Further analysis regarding the difference in the VPg sequence of the predominant strain of BaYMV collected from these two testing sites may explain the difference of resistance loci differentially identified under geographically distinct regions. Our research provides novel genetic resources and resistance loci for breeding barley varieties for BaMYV disease resistance.

Genomic and Pathogenic Diversity of Barley Yellow Mosaic Virus and Barley Mild Mosaic Virus Isolates in Fields of China and Their Compatibility with Resistance Genes of Cultivated Barley.

Plant viruses transmitted by the soilborne plasmodiophorid Polymyxa graminis constantly threaten global production of cereal crops. Although the yellow mosaic virus disease of barley has been known to be present for a long time in China, the understanding of the diversity of the viral pathogens and their interactions with host resistance remains limited. In this study, we conducted a nationwide survey of P. graminis and the barley yellow mosaic virus (BaYMV) and barley mild mosaic virus (BaMMV) it transmits, followed by genomic and pathogenic diversity analyses of both viruses. BaYMV and BaMMV were found exclusively in the region downstream of the Yangtze River, despite the national distribution of its transmission vector P. graminis. Analysis of the genomic variations of BaYMV and BaMMV revealed an elevated rate of nonsynonymous substitutions in the viral genome-linked protein (VPg), in which most substitutions were located in its interaction surface with the host eukaryotic translation initiation factor 4E (eIF4E). VPg sequence diversity was associated with the divergence in virus pathogenicity that was identified through multiple field trials. The majority of the resistance genes, including the widely applied rym4 and rym5 (alleles of eIF4E), as well as the combination of rym1/11 and rym5, are not sufficient to protect cultivated barley against viruses in China. Collectively, these results provide insights into virulence specificity and interaction mode with host resistance in cultivated barley, which has significant implications in breeding for the broad-spectrum resistance barley varieties.

Identification of novel QTL contributing to barley yellow mosaic resistance in wild barley (Hordeum vulgare spp. spontaneum).

BACKGROUND: Barley yellow mosaic disease (BYMD) caused by Barley yellow mosaic virus (BaYMV) and Barley mild mosaic virus (BaMMV) seriously threatens the production of winter barley. Cultivating and promoting varieties that carry disease-resistant genes is one of the most powerful ways to minimize the disease's effect on yield. However, as the BYMD virus mutates rapidly, resistance conferred by the two cloned R genes to the virus had been overcome by new virus strains. There is an urgent need for novel resistance genes in barley that convey sustainable resistance to newly emerging virus strains causing BYMD. RESULTS: A doubled haploid (DH) population derived from a cross of SRY01 (BYMD resistant wild barley) and Gairdner (BYMD susceptible barley cultivar) was used to explore for QTL of resistance to BYMD in barley. A total of six quantitative trait loci (qRYM-1H, qRYM-2Ha, qRYM-2Hb, qRYM-3H, qRYM-5H, and qRYM-7H) related to BYMD resistance were detected, which were located on chromosomes 1H, 2H, 3H, 5H, and 7H. Both qRYM-1H and qRYM-2Ha were detected in all environments. qRYM-1H was found to be overlapped with rym7, a known R gene to the disease, whereas qRYM-2Ha is a novel QTL on chromosome 2H originated from SRY01, explaining phenotypic variation from 9.8 to 17.8%. The closely linked InDel markers for qRYM-2Ha were developed which could be used for marker-assisted selection in barley breeding. qRYM-2Hb and qRYM-3H were stable QTL for specific resistance to Yancheng and Yangzhou virus strains, respectively. qRYM-5H and qRYM-7H identified in Yangzhou were originated from Gairdner. CONCLUSIONS: Our work is focusing on a virus disease (barley yellow mosaic) of barley. It is the first report on BYMD-resistant QTL from wild barley accessions. One novel major QTL (qRYM-2Ha) for the resistance was detected. The consistently detected new genes will potentially serve as novel sources for achieving pre-breeding barley materials with resistance to BYMD.

Rapid Generation and Analysis of a Barley Doubled Haploid Line with Higher Nitrogen Use Efficiency Than Parental Lines by F1 Microspore Embryogenesis.

Creating varieties with high nitrogen use efficiency (NUE) is crucial for sustainable agriculture development. In this study, a superior barley doubled haploid line (named DH45) with improved NUE was produced via F1 microspore embryogenesis with three rounds of screening in different nitrogen levels by hydroponic and field experiments. The molecular mechanisms responsible for the NUE of DH45 surpassing that of its parents were investigated by RNA-seq analysis. A total of 1027 differentially expressed genes (DEGs) were identified that were up- or down-regulated in DH45 under low nitrogen conditions but showed no significant differences in the parents. GO analysis indicated that genes involved in nitrogen compound metabolic processes were significantly enriched in DH45 compared with the parents. KEGG analysis showed the MAPK signaling pathway plant to be highly enriched in DH45 relative to its parents, as well as genes involved in alanine, aspartate and glutamate metabolism, and arginine biosynthesis. In conclusion, our study revealed the potential to fix trait superiority in a line by combining crossing with F1 microspore culture technologies in future crop breeding and also identified several candidate genes that are expressed in shoots and may enable barley to cope with low-nitrogen stress.

Characterization of the Nitrate Transporter gene family and functional identification of HvNRT2.1 in barley (Hordeum vulgare L.).

Nitrogen use efficiency (NUE) is the efficiency with which plants acquire and use nitrogen. Plants have high-affinity nitrate transport systems, which involve certain nitrate transporter (NRT) genes. However, limited data are available on the contribution of the NRT2/3 gene family in barley nitrate transport. In the present study, ten putative NRT2 and three putative NRT3 genes were identified using bioinformatics methods. All the HvNRT2/3 genes were located on chromosomes 3H, 5H, 6H or 7H. Remarkably, the presence of tandem repeats indicated that duplication events contributed to the expansion of the NRT2 gene family in barley. In addition, the HvNRT2/3 genes displayed various expression patterns at selected developmental stages and were induced in the roots by both low and high nitrogen levels. Furthermore, the overexpression of HvNRT2.1 improved the yield related traits in Arabidopsis. Taken together, the data generated in the present study will be useful for genome-wide analyses to determine the precise role of the HvNRT2/3 genes during barley development, with the ultimate goal of improving NUE and crop production.

Overexpression of Barley Transcription Factor HvERF2.11 in Arabidopsis Enhances Plant Waterlogging Tolerance.

Waterlogging stress significantly affects the growth, development, and productivity of crop plants. However, manipulation of gene expression to enhance waterlogging tolerance is very limited. In this study, we identified an ethylene-responsive factor from barley, which was strongly induced by waterlogging stress. This transcription factor named HvERF2.11 was 1158 bp in length and encoded 385 amino acids, and mainly expressed in the adventitious root and seminal root. Overexpression of HvERF2.11 in Arabidopsis led to enhanced tolerance to waterlogging stress. Further analysis of the transgenic plants showed that the expression of AtSOD1, AtPOD1 and AtACO1 increased rapidly, while the same genes did not do so in non-transgenic plants, under waterlogging stress. Activities of antioxidant enzymes and alcohol dehydrogenase (ADH) were also significantly higher in the transgenic plants than in the non-transgenic plants under waterlogging stress. Therefore, these results indicate that HvERF2.11 plays a positive regulatory role in plant waterlogging tolerance through regulation of waterlogging-related genes, improving antioxidant and ADH enzymes activities.

Understanding Mechanisms of Salinity Tolerance in Barley by Proteomic and Biochemical Analysis of Near-Isogenic Lines.

Salt stress is one of the major environmental factors impairing crop production. In our previous study, we identified a major QTL for salinity tolerance on chromosome 2H on barley (Hordeum vulgare L.). For further investigation of the mechanisms responsible for this QTL, two pairs of near-isogenic lines (NILs) differing in this QTL were developed. Sensitive NILs (N33 and N53) showed more severe damage after exposure to 300 mM NaCl than tolerant ones (T46 and T66). Both tolerant NILs maintained significantly lower Na+ content in leaves and much higher K+ content in the roots than sensitive lines under salt conditions, thus indicating the presence of a more optimal Na+/K+ ratio in plant tissues. Salinity stress caused significant accumulation of H2O2, MDA, and proline in salinity-sensitive NILs, and a greater enhancement in antioxidant enzymatic activities at one specific time or tissues in tolerant lines. One pair of NILs (N33 and T46) were used for proteomic studies using two-dimensional gel electrophoresis. A total of 53 and 51 differentially expressed proteins were identified through tandem mass spectrometry analysis in the leaves and roots, respectively. Proteins which are associated with photosynthesis, reactive oxygen species (ROS) scavenging, and ATP synthase were found to be specifically upregulated in the tolerant NIL. Proteins identified in this study can serve as a useful resource with which to explore novel candidate genes for salinity tolerance in barley.

Genome wide association study reveals novel QTL for barley yellow dwarf virus resistance in wheat.

BACKGROUND: Barley yellow dwarf (BYD) is an important virus disease that causes significant reductions in wheat yield. For effective control of Barley yellow dwarf virus through breeding, the identification of genetic sources of resistance is key to success. In this study, 335 geographically diverse wheat accessions genotyped using an Illumina iSelect 90 K single nucleotide polymorphisms (SNPs) bead chip array were used to identify new sources of resistance to BYD in different environments. RESULTS: A genome-wide association study (GWAS) performed using all the generalised and mixed linkage models (GLM and MLM, respectively) identified a total of 36 significant marker-trait associations, four of which were consistently detected in the K model. These four novel quantitative trait loci (QTL) were identified on chromosomes 2A, 2B, 6A and 7A and associated with markers IWA3520, IWB24938, WB69770 and IWB57703, respectively. These four QTL showed an additive effect with the average visual symptom score of the lines containing resistance alleles of all four QTL being much lower than those with less favorable alleles. Several Chinese landraces, such as H-205 (Baimazha) and H-014 (Dahongmai) which have all four favorable alleles, showed consistently higher resistance in different field trials. None of them contained the previously described Bdv2, Bdv3 or Bdv4 genes for BYD resistance. CONCLUSIONS: This study identified multiple novel QTL for BYD resistance and some resistant wheat genotypes. These will be useful for breeders to generate combinations with and/or without Bdv2 to achieve higher levels and more stable BYD resistance.

Regulation of nitrogen availability results in changes in grain protein content and grain storage subproteomes in barley (Hordeum vulgare L.).

Barley grain protein content (GPC) is an important quality factor that determines grain end-use value. The synthesis and accumulation of grain protein is highly dependent on the availability of nitrogen fertilizer, and it is important to understand the underlying control mechanisms of this. In the current study, the GPC and protein composition of mature grain seeds from Yangsimai 3 and Naso Nijo barley cultivars were analyzed. Grain storage subproteomes (albumin, glubulin, hordein and glutelin) were compared in the cultivars grown in both low and high nitrogen level conditions. The GPC of mature grain was significantly higher in Yangsimai 3 than Naso Nijo following nitrogen treatment. Albumin, hordein and glutelin content were increased in Yangsimai, while only hordein content was increased in Naso Nijo. Large-scale analysis of the grain storage subproteome revealed 152 differentially expressed protein spots on 2-DE gels with a pH range of 3-10. Among these, 42 and 66 protein spots were successfully identified by tandem mass spectrometry in Yangsimai 3 and Naso Nijo grown in low and high nitrogen conditions. The identified proteins were further grouped into thirteen categories according to their biological functions. This detailed analysis of grain subproteomes provides information on how barley GPC may be controlled by nitrogen supply.