Identification of a novel and plant height-independent QTL for coleoptile length in barley and validation of its effect using near isogenic lines.

KEY MESSAGE: This study reported the identification and validation of novel QTL conferring coleoptile length in barley and predicted candidate genes underlying the largest effect QTL based on orthologous analysis and comparison of the whole genome assemblies for both parental genotypes of the mapping population. Coleoptile length (CL) is one of the most important agronomic traits in cereal crops due to its direct influence on the optimal depth for seed sowing which facilitates better seedling establishment. Varieties with longer coleoptiles are preferred in drought-prone areas where less moisture maintains at the top layer of the soil. Compared to wheat, genetic study on coleoptile length is limited in barley. Here, we reported a study on detecting the genomic regions associated with CL in barley by assessing a population consisting of 201 recombinant inbred lines. Four putative QTL conferring CL were consistently identified on chromosomes 1H, 5H, 6H, and 7H in each of the trials conducted. Of these QTL, the two located on chromosomes 5H and 6H (designated as Qcl.caf-5H and Qcl.caf-6H) are likely novel and Qcl.caf-5H showed the most significant effect explaining up to 30.9% of phenotypic variance with a LOD value of 15.1. To further validate the effect of this putative QTL, five pairs of near isogenic lines (NILs) were then developed and assessed. Analysis of the NILs showed an average difference of 21.0% in CL between the two isolines. Notably, none of the other assessed morphological characteristics showed consistent differences between the two isolines for each pair of the NILs. Candidate genes underlying the Qcl.caf-5H locus were also predicted by employing orthologous analysis and comparing the genome assemblies for both parental genotypes of the mapping population in the present study. Taken together, these findings expand our understanding on genetic basis of CL and will be indicative for further gene cloning and functional analysis underly this locus in barley.

Transcriptomic insights into shared responses to Fusarium crown rot infection and drought stresses in bread wheat (Triticum aestivum L.).

KEY MESSAGE: Shared changes in transcriptomes caused by Fusarium crown rot infection and drought stress were investigated based on a single pair of near-isogenic lines developed for a major locus conferring tolerance to both stresses. Fusarium crown rot (FCR) is a devastating disease in many areas of cereal production worldwide. It is well-known that drought stress enhances FCR severity but possible molecular relationship between these two stresses remains unclear. To investigate their relationships, we generated several pairs of near isogenic lines (NILs) targeting a locus conferring FCR resistance on chromosome 2D in bread wheat. One pair of these NILs showing significant differences between the two isolines for both FCR resistance and drought tolerance was used to investigate transcriptomic changes in responsive to these two stresses. Our results showed that the two isolines likely deployed different strategies in dealing with the stresses, and significant differences in expressed gene networks exist between the two time points of drought stresses evaluated in this study. Nevertheless, results from analysing Gene Ontology terms and transcription factors revealed that similar regulatory frameworks were activated in coping with these two stresses. Based on the position of the targeted locus, changes in expression following FCR infection and drought stresses, and the presence of non-synonymous variants between the two isolines, several candidate genes conferring resistance or tolerance to these two types of stresses were identified. The NILs generated, the large number of DEGs with single-nucleotide polymorphisms detected between the two isolines, and the candidate genes identified would be invaluable in fine mapping and cloning the gene(s) underlying the targeted locus.

A co-located QTL for seven spike architecture-related traits shows promising breeding use potential in common wheat (Triticum aestivum L.).

KEY MESSAGE: A co-located novel QTL for TFS, FPs, FMs, FFS, FFPs, KWS, and KWPs with potential of improving wheat yield was identified and validated. Spike-related traits, including fertile florets per spike (FFS), kernel weight per spike (KWS), total florets per spike (TFS), florets per spikelet (FPs), florets in the middle spikelet (FMs), fertile florets per spikelet (FFPs), and kernel weight per spikelet (KWPs), are key traits in improving wheat yield. In the present study, quantitative trait loci (QTL) for these traits evaluated under various environments were detected in a recombinant inbred line population (msf/Chuannong 16) mainly genotyped using the 16 K SNP array. Ultimately, we identified 60 QTL, but only QFFS.sau-MC-1A for FFS was a major and stably expressed QTL. It was located on chromosome arm 1AS, where loci for TFS, FPs, FMs, FFS, FFPs, KWS, and KWPs were also simultaneously co-mapped. The effect of QFFS.sau-MC-1A was further validated in three independent segregating populations using a Kompetitive Allele-Specific PCR marker. For the co-located QTL, QFFS.sau-MC-1A, the presence of a positive allele from msf was associate with increases for all traits: + 12.29% TFS, + 10.15% FPs, + 13.97% FMs, + 17.12% FFS, + 14.75% FFPs, + 22.17% KWS, and + 19.42% KWPs. Furthermore, pleiotropy analysis showed that the positive allele at QFFS.sau-MC-1A simultaneously increased the spike length, spikelet number per spike, and thousand-kernel weight. QFFS.sau-MC-1A represents a novel QTL for marker-assisted selection with the potential for improving wheat yield. Four genes, TraesCS1A03G0012700, TraesCS1A03G0015700, TraesCS1A03G0016000, and TraesCS1A03G0016300, which may affect spike development, were predicted in the physical interval harboring QFFS.sau-MC-1A. Our results will help in further fine mapping QFFS.sau-MC-1A and be useful for improving wheat yield.

Fine mapping of a Fusarium crown rot resistant locus on chromosome arm 6HL in barley by exploiting near isogenic lines, transcriptome profiling, and a large near isogenic line-derived population.

KEY MESSAGE: This study reported validation and fine mapping of a Fusarium crown rot resistant locus on chromosome arm 6HL in barley using near isogenic lines, transcriptome sequences, and a large near isogenic line-derived population. Fusarium crown rot (FCR), caused by Fusarium pseudograminearum, is a chronic and serious disease affecting cereal production in semi-arid regions globally. The increasing prevalence of this disease in recent years is attributed to the widespread adoption of minimum tillage and stubble retention practices. In the study reported here, we generated eight pairs of near isogenic lines (NILs) targeting a putative QTL (Qcrs.caf-6H) conferring FCR resistance in barley. Assessing the NILs confirmed the large effect of this locus. Aimed to develop markers that can be reliably used in incorporating this resistant allele into breeding programs and identify candidate genes, transcriptomic analyses were conducted against three of the NIL pairs and a large NIL-derived population consisting of 1085 F7 recombinant inbred lines generated. By analyzing the transcriptomic data and the fine mapping population, Qcrs.caf-6H was delineated into an interval of 0.9 cM covering a physical distance of ~ 547 kb. Six markers co-segregating with this locus were developed. Based on differential gene expression and SNP variations between the two isolines among the three NIL pairs, candidate genes underlying the resistance at this locus were detected. These results would improve the efficiency of incorporating the targeted locus into barley breeding programs and facilitate the cloning of causal gene(s) responsible for the resistance.

A major QTL simultaneously increases the number of spikelets per spike and thousand-kernel weight in a wheat line.

KEY MESSAGE: A novel and stably expressed QTL QSNS.sicau-SSY-7A for spikelet number per spike in wheat without negative effects on thousand-kernel weight was identified and validated in different genetic backgrounds. Spikelet number per spike (SNS) is an important determinant of yield in wheat. In the present study, we combined bulked segregant analysis (BSA) and the wheat 660 K single-nucleotide polymorphism (SNP) array to rapidly identify genomic regions associated with SNS from a recombinant inbred line (RIL) population derived from a cross between the wheat lines S849-8 and SY95-71. A genetic map was constructed using Kompetitive Allele Specific PCR markers in the SNP-enriched region on the long arm of chromosome 7A. A major and stably expressed QTL, QSNS.sicau-SSY-7A, was detected in multiple environments. It was located in a 1.6 cM interval on chromosome arm 7AL flanked by the markers AX-109983514 and AX-109820548. This QTL explained 6.86-15.72% of the phenotypic variance, with LOD values ranging from 3.66 to 8.66. Several genes associated with plant growth and development were identified in the interval where QSNS.sicau-SSY-7A was located on the 'Chinese Spring' wheat and wild emmer reference genomes. Furthermore, the effects of QSNS.sicau-SSY-7A and WHEAT ORTHOLOG OFAPO1(WAPO1) on SNS were analyzed. Interestingly, QSNS.sicau-SSY-7A significantly increased SNS without negative effects on thousand-kernel weight, anthesis date and plant height, demonstrating its great potential for breeding aimed at improving grain yield. Taken together, these results indicate that QSNS.sicau-SSY-7A is a promising locus for yield improvement, and its linkage markers are helpful for fine mapping and molecular breeding.

Does one subgenome become dominant in the formation and evolution of a polyploid?

BACKGROUND: Polyploids are common in flowering plants and they tend to have more expanded ranges of distributions than their diploid progenitors. Possible mechanisms underlying polyploid success have been intensively investigated. Previous studies showed that polyploidy generates novel changes and that subgenomes in allopolyploid species often differ in gene number, gene expression levels and levels of epigenetic alteration. It is widely believed that such differences are the results of conflicts among the subgenomes. These differences have been treated by some as subgenome dominance, and it is claimed that the magnitude of subgenome dominance increases in polyploid evolution. SCOPE: In addition to changes which occurred during evolution, differences between subgenomes of a polyploid species may also be affected by differences between the diploid donors and changes which occurred during polyploidization. The variable genome components in many plant species are extensive, which would result in exaggerated differences between a subgenome and its progenitor when a single genotype or a small number of genotypes are used to represent a polyploid or its donors. When artificially resynthesized polyploids are used as surrogates for newly formed genotypes which have not been exposed to evolutionary selection, differences between diploid genotypes available today and those involved in the formation of the natural polyploid genotypes must also be considered. CONCLUSIONS: Contrary to the now widely held views that subgenome biases in polyploids are the results of conflicts among the subgenomes and that one of the parental subgenomes generally retains more genes which are more highly expressed, available results show that subgenome biases mainly reflect legacy from the progenitors and that they can be detected before the completion of polyploidization events. Further, there is no convincing evidence that the magnitudes of subgenome biases have significantly changed during evolution for any of the allopolyploid species assessed.

A new major QTL for flag leaf thickness in barley (Hordeum vulgare L.).

BACKGROUND: Carbohydrate accumulation of photosynthetic organs, mainly leaves, are the primary sources of grain yield in cereals. The flag leaf plays a vital role in seed development, which is probably the most neglected morphological characteristic during traditional selection processes. RESULTS: In this experiment, four flag leaf morphological traits and seven yield-related traits were investigated in a DH population derived from a cross between a wild barley and an Australian malting barley cultivar. Flag leaf thickness (FLT) showed significantly positive correlations with grain size. Four QTL, located on chromosomes 1H, 2H, 3H, and 5H, respectively, were identified for FLT. Among them, a major QTL was located on chromosome 3H with a LOD value of 18.4 and determined 32% of the phenotypic variation. This QTL showed close links but not pleiotropism to the previously reported semi-dwarf gene sdw1 from the cultivated barley. This QTL was not reported before and the thick leaf allele from the wild barley could provide a useful source for improving grain yield through breeding. CONCLUSIONS: Our results also provided valuable evidence that source traits and sink traits in barley are tightly connected and suggest further improvement of barley yield potential with enhanced and balanced source and sink relationships by exploiting potentialities of the wild barley resources. Moreover, this study will provide a novel sight on understanding the evolution and development of leaf morphology in barley and improving barley production by rewilding for lost superior traits during plant evolution.

Leaf thickness of barley: genetic dissection, candidate genes prediction and its relationship with yield-related traits.

KEY MESSAGE: In this first genetic study on assessing leaf thickness directly in cereals, major and environmentally stable QTL were detected in barley and candidate genes underlying a major locus were identified. Leaf thickness (LT) is an important characteristic affecting leaf functions which have been intensively studied. However, as LT has a small dimension in many plant species and technically difficult to measure, previous studies on this characteristic are often based on indirect estimations. In the first study of detecting QTL controlling LT by directly measuring the characteristic in barley, large and stable loci were detected from both field and glasshouse trials conducted in different cropping seasons by assessing a population of 201 recombinant inbred lines. Four loci (locating on chromosome arms 2H, 3H, 5H and 6H, respectively) were consistently detected for flag leaf thickness (FLT) in each of these trials. The one on 6H had the largest effect, with a maximum LOD 9.8 explaining up to 20.9% of phenotypic variance. FLT does not only show strong interactions with flag leaf width and flag leaf area but has also strong correlations with fertile tiller number, spike row types, kernel number per spike and heading date. Though with reduced efficiency, these loci were also detectable from assessing second last leaf of fully grown plants or even from assessing the third leaves of seedlings. Taking advantage of the high-quality genome assemblies for both parents of the mapping population used in this study, three candidate genes underlying the 6H QTL were predicted based on orthologous analysis. These results do not only broaden our understanding on genetic basis of LT and its relationship with other traits in cereal crops but also form the bases for cloning and functional analysis of genes regulating LT in barley.

Differences between diploid donors are the main contributing factor for subgenome asymmetry measured in either gene ratio or relative diversity in allopolyploids.

Subgenome asymmetry (SA) has routinely been attributed to different responses between the subgenomes of a polyploid to various stimuli during evolution. Here, we compared subgenome differences in gene ratio and relative diversity between artificial and natural genotypes of several allopolyploid species. Surprisingly, consistent differences were not detected between these two types of polyploid genotypes, although they differ in times exposed to evolutionary selection. The estimated ratio of shared genes between a subgenome and its diploid donor was invariably higher for the artificial allopolyploid genotypes than those for the natural genotypes, which is expected as it is now well-known that many genes in a species are not shared among all individuals. As the exact diploid parent for a given subgenome is unknown, the estimated ratios of shared genes for the natural genotypes would also include difference among individual genotypes of the diploid donor species. Further, we detected the presence of SA in genotypes before the completion of the polyploidization events as well as in those which were not formed via polyploidization. These results indicate that SA may, to a large degree, reflect differences between its diploid donors or that changes occurred during polyploid evolution are defined by their donor genomes.

Detection of a major QTL conditioning trichome length and density on chromosome arm 4BL and development of near isogenic lines targeting this locus in bread wheat.

Trichomes are differentiated epidermal cells and can be found on above ground organs of nearly all land plants. Results from previous studies show that trichomes play important roles against a wide range of both biotic and abiotic stresses. By examining differences between parental genotypes of available populations, we identified a population of recombinant inbred lines showing clear segregation for trichome density and length. Assessing the F8 lines of the population growing in the field detected a major locus on chromosome arm 4BL. This locus was detected based the assessments of either fully expanded third leaves or flag leaves after anthesis. Based on the position of the QTL, an SSR marker was used to identify heterozygous plants at this locus from F5 lines derived from the same cross for the F8 population. Three pairs of near isogenic lines targeting this locus were obtained from these heterozygous plants. Difference in trichome length between the two lines with opposite alleles for each of these NIL pairs were similar to that between the two parental genotypes for the mapping populations, confirming that this single locus is mainly responsible for the trichome characteristics measured in this study. The allele with long and dense trichome is dominant as this characteristic was shown by the heterozygous individuals at this marker locus. Apart from the targeted locus, NIL pairs have highly homogeneous genetic backgrounds. Thus, the NILs could be invaluable in understanding the relationship between trichome density and resistance or tolerance to various biotic and abiotic stresses. Supplementary Information: The online version contains supplementary material available at 10.1007/s11032-021-01201-8.

The draft genome of a wild barley genotype reveals its enrichment in genes related to biotic and abiotic stresses compared to cultivated barley.

Wild barley (Hordeum spontaneum) is the progenitor of cultivated barley (Hordeum vulgare) and provides a rich source of genetic variations for barley improvement. Currently, the genome sequences of wild barley and its differences with cultivated barley remain unclear. In this study, we report a high-quality draft assembly of wild barley accession (AWCS276; henceforth named as WB1), which consists of 4.28 Gb genome and 36 395 high-confidence protein-coding genes. BUSCO analysis revealed that the assembly included full lengths of 95.3% of the 956 single-copy plant genes, illustrating that the gene-containing regions have been well assembled. By comparing with the genome of the cultivated genotype Morex, it is inferred that the WB1 genome contains more genes involved in resistance and tolerance to biotic and abiotic stresses. The presence of the numerous WB1-specific genes indicates that, in addition to enhance allele diversity for genes already existing in the cultigen, exploiting the wild barley taxon in breeding should also allow the incorporation of novel genes. Furthermore, high levels of genetic variation in the pericentromeric regions were detected in chromosomes 3H and 5H between the wild and cultivated genotypes, which may be the results of domestication. This H. spontaneum draft genome assembly will help to accelerate wild barley research and be an invaluable resource for barley improvement and comparative genomics research.

Identifying barley pan-genome sequence anchors using genetic mapping and machine learning.

KEY MESSAGE: We identified 1.844 million barley pan-genome sequence anchors from 12,306 genotypes using genetic mapping and machine learning. There is increasing evidence that genes from a given crop genotype are far to cover all genes in that species; thus, building more comprehensive pan-genomes is of great importance in genetic research and breeding. Obtaining a thousand-genotype scale pan-genome using deep-sequencing data is currently impractical for species like barley which has a huge and highly repetitive genome. To this end, we attempted to identify barley pan-genome sequence anchors from a large quantity of genotype-by-sequencing (GBS) datasets by combining genetic mapping and machine learning algorithms. Based on the GBS sequences from 11,166 domesticated and 1140 wild barley genotypes, we identified 1.844 million pan-genome sequence anchors. Of them, 532,253 were identified as presence/absence variation (PAV) tags. Through aligning these PAV tags to the genome of hulless barley genotype Zangqing320, our analysis resulted in a validation of 83.6% of them from the domesticated genotypes and 88.6% from the wild barley genotypes. Association analyses against flowering time, plant height and kernel size showed that the relative importance of the PAV and non-PAV tags varied for different traits. The pan-genome sequence anchors based on GBS tags can facilitate the construction of a comprehensive pan-genome and greatly assist various genetic studies including identification of structural variation, genetic mapping and breeding in barley.

A pan-transcriptome analysis shows that disease resistance genes have undergone more selection pressure during barley domestication.

BACKGROUND: It has become clear in recent years that many genes in a given species may not be found in a single genotype thus using sequences from a single genotype as reference may not be adequate for various applications. RESULTS: In this study we constructed a pan-transcriptome for barley by de novo assembling 288 sets of RNA-seq data from 32 cultivated barley genotypes and 31 wild barley genotypes. The pan-transcriptome consists of 756,632 transcripts with an average N50 length of 1240 bp. Of these, 289,697 (38.2%) were not found in the genome of the international reference genotype Morex. The novel transcripts are enriched with genes associated with responses to different stresses and stimuli. At the pan-transcriptome level, genotypes of wild barley have a higher proportion of disease resistance genes than cultivated ones. CONCLUSIONS: We demonstrate that the use of the pan-transcriptome dramatically improved the efficiency in detecting variation in barley. Analysing the pan-transcriptome also found that, compared with those in other categories, disease resistance genes have gone through stronger selective pressures during domestication.

Validation and delineation of a locus conferring Fusarium crown rot resistance on 1HL in barley by analysing transcriptomes from multiple pairs of near isogenic lines.

BACKGROUND: Fusarium crown rot (FCR) is a chronic and severe disease in cereal production in semi-arid regions worldwide. A putative quantitative trait locus conferring FCR resistance, Qcrs.cpi-1H, had previously been mapped on the long arm of chromosome 1H in barley. RESULTS: In this study, five pairs of near-isogenic lines (NILs) targeting the 1HL locus were developed. Analysing the NILs found that the resistant allele at Qcrs.cpi-1H significantly reduced FCR severity. Transcriptomic analysis was then conducted against three of the NIL pairs, which placed the Qcrs.cpi-1H locus in an interval spanning about 11 Mbp. A total of 56 expressed genes bearing single nucleotide polymorphisms (SNPs) were detected in this interval. Five of them contain non-synonymous SNPs. These results would facilitate detailed mapping as well as cloning gene(s) underlying the resistance locus. CONCLUSION: NILs developed in this study and the transcriptomic sequences obtained from them did not only allow the validation of the resistance locus Qcrs.cpi-1H and the identification of candidate genes underlying its resistance, they also allowed the delineation of the resistance locus and the development of SNPs markers which formed a solid base for detailed mapping as well as cloning gene(s) underlying the locus.

Genome-wide association study reveals new loci for yield-related traits in Sichuan wheat germplasm under stripe rust stress.

BACKGROUND: As one of the most important food crops in the world, increasing wheat (Triticum aestivum L.) yield is an urgent task for global food security under the continuous threat of stripe rust (caused by Puccinia striiformis f. sp. tritici) in many regions of the world. Molecular marker-assisted breeding is one of the most efficient ways to increase yield. Here, we identified loci associated to multi-environmental yield-related traits under stripe rust stress in 244 wheat accessions from Sichuan Province through genome-wide association study (GWAS) using 44,059 polymorphic markers from the 55 K single nucleotide polymorphism (SNP) chip. RESULTS: A total of 13 stable quantitative trait loci (QTLs) were found to be highly associating to yield-related traits, including 6 for spike length (SL), 3 for thousand-kernel weight (TKW), 2 for kernel weight per spike (KWPS), and 2 for both TKW and KWPS, in at least two test environments under stripe rust stress conditions. Of them, ten QTLs were overlapped or very close to the reported QTLs, three QTLs, QSL.sicau-1AL, QTKW.sicau-4AL, and QKWPS.sicau-4AL.1, were potentially novel through the physical location comparison with previous QTLs. Further, 21 candidate genes within three potentially novel QTLs were identified, they were mainly involved in the regulation of phytohormone, cell division and proliferation, meristem development, plant or organ development, and carbohydrate transport. CONCLUSIONS: QTLs and candidate genes detected in our study for yield-related traits under stripe rust stress will facilitate elucidating genetic basis of yield-related trait and could be used in marker-assisted selection in wheat yield breeding.

Development of tightly linked markers and identification of candidate genes for Fusarium crown rot resistance in barley by exploiting a near-isogenic line-derived population.

KEY MESSAGE: This study demonstrates the feasibility of developing co-segregating markers and identifying candidate genes for Fusarium crown rot resistance in barley based on the generation and exploitation of a near-isogenic line-derived large population. Fusarium crown rot (FCR) is a chronic and severe disease in cereals in semi-arid regions worldwide. Previous studies showed that FCR assessment could be affected by many factors including plant height, growth rate as well as drought stress. Thus, accurate assessment, which is essential for detailed mapping of any locus conferring FCR resistance, is difficult. Targeting one of the resistance loci reported earlier, we developed a near-isogenic line-derived population consisting of 1820 F9 lines. By analysing this population, the Qcrs.cpi-4H locus was mapped to an interval of 0.09 cM covering a physical distance of about 637 kb and 13 markers co-segregating with the targeted locus were developed. Candidate genes underlying the resistance locus were identified by analysing the expression and sequence variation of genes in the targeted interval. The accurate localization and the development of co-segregating markers should facilitate the incorporation of this large-effect QTL into breeding programmes as well as the cloning of gene(s) underlying the locus.

Identification and validation of a novel major QTL for all-stage stripe rust resistance on 1BL in the winter wheat line 20828.

KEY MESSAGE: A major, likely novel stripe rust resistance QTL for all-stage resistance on chromosome arm 1BL identified in a 1.76-cM interval using a saturated linkage map was validated in four populations with different genetic backgrounds. Stripe rust is a globally important disease of wheat. Identification and utilization of new resistance genes are essential for breeding resistant cultivars. Wheat line 20828 has exhibited high levels of stripe rust resistance for over a decade. However, the genetics of stripe rust resistance in this line has not been studied. A set of 199 recombinant inbred lines (RILs) were developed from a cross between 20828 and a susceptible cultivar Chuannong 16. The RIL population was genotyped with the Wheat55K SNP (single nucleotide polymorphism) array and SSR (simple sequence repeat) markers and evaluated in four environments with current predominant Puccinia striiformis f. sp. tritici t races including CYR32, CYR33 and CYR34. Four stable QTL were located on chromosomes 1B (2 QTL), 4A and 6A. Among them, the major QTL, QYr.sicau-1B.1 (LOD = 23-28, PVE = 16-39%), was localized to a 1.76-cM interval flanked by SSR markers Xwmc216 and Xwmc156 on chromosome 1BL. Eight resistance genes were previously identified in the physical interval of QYr.sicau-1B.1. Compared with previous studies, QYr.sicau-1B.1 is a new gene for resistant to stripe rust. It was further verified by analysis of the closely linked SSR markers Xwmc216 and Xwmc156 in four other populations with different genetic backgrounds. QYr.sicau-1B.1 reduced the stripe rust disease index by up to 82.8%. Three minor stable QTL (located on chromosomes 1B, 4A and 6A, respectively) also added to the resistance level of QYr.sicau-1B.1. Our results provide valuable information for further fine mapping and cloning as well as molecular-assisted breeding with QYr.sicau-1B.1.

A multiple near isogenic line (multi-NIL) RNA-seq approach to identify candidate genes underpinning QTL.

KEY MESSAGE: This study demonstrates how identification of genes underpinning disease-resistance QTL based on differential expression and SNPs can be improved by performing transcriptomic analysis on multiple near isogenic lines. Transcriptomic analysis has been widely used to understand the genetic basis of a trait of interest by comparing genotypes with contrasting phenotypes. However, these approaches identify such large sets of differentially expressed genes that it proves difficult to isolate which genes underpin the phenotype of interest. This study tests whether using multiple near isogenic lines (NILs) can improve the resolution of RNA-seq-based approaches to identify genes underpinning disease-resistance QTL. A set of NILs for a major effect Fusarium crown rot-resistance QTL in barley on the 4HL chromosome arm were analysed under Fusarium crown rot using RNA-seq. Differential gene expression and single nucleotide polymorphism detection analyses reduced the number of putative candidates from thousands within individual NIL pairs to only one hundred and two genes, which were differentially expressed or contained SNPs in common across NIL pairs and occurred on 4HL. Our findings support the value of performing RNA-seq analysis using multiple NILs to remove genetic background effects. The enrichment analyses indicated conserved differences in the response to infection between resistant and sensitive isolines suggesting that sensitive isolines are impaired in systemic defence response to Fusarium pseudograminearum.

Sequence divergence between spelt and common wheat.

KEY MESSAGE: Sequence comparison between spelt and common wheat reveals that the former has huge potential in enriching the genetic variation of the latter. Genetic variation is the foundation of crop improvement. By comparing genome sequences of a Triticum spelta accession and one of its derived hexaploid lines with the sequences of the international reference genotype Chinese Spring, we detected variants more than tenfold higher than those present among common wheat (T. aestivum L) genotypes. Furthermore, different from the typical 'V-shaped' pattern of variant distribution often observed along wheat chromosomes, the sequence variation detected in this study was more evenly distributed along the 3B chromosome. This was also the case between T. spelta and the wild emmer genome. Genetic analysis showed that T. spelta and common wheat formed discrete groups. These results showed that, although it is believed that the spelt and common wheat are evolutionarily closely related and belong to the same species, a significant sequence divergence exists between them. Thus, the values of T. spelta in enriching the genetic variation of common wheat can be huge.

A 55 K SNP array-based genetic map and its utilization in QTL mapping for productive tiller number in common wheat.

KEY MESSAGE: A high-density genetic map constructed with a wheat 55 K SNP array was highly consistent with the physical map of this species and it facilitated the identification of a novel major QTL for productive tiller number. Productive tiller number (PTN) plays a key role in wheat grain yield. In this study, a recombinant inbred line population with 199 lines derived from a cross between '20828' and 'Chuannong16' was used to construct a high-density genetic map using wheat 55 K single nucleotide polymorphism (SNP) array. The constructed genetic map contains 12,109 SNP markers spanning 3021.04 cM across the 21 wheat chromosomes. The orders of the genetic and physical positions of these markers are generally in agreement, and they also match well with those based on the 660 K SNP array from which the one used in this study was derived. The ratios of SNPs located in each of the wheat deletion bins were similar among the wheat 9 K, 55 K, 90 K, 660 K and 820 K SNP arrays. Based on the constructed maps, a novel major quantitative trait locus QPtn.sau-4B for PTN was detected across multi-environments in a 0.55 cM interval on 4B and it explained 17.23-45.46% of the phenotypic variance. Twenty common genes in the physical interval between the flanking markers were identified on chromosome 4B of 'Chinese Spring' and wild emmer. These results indicate that wheat 55 K SNP array could be an ideal tool in primary mapping of target genes and the identification of QPtn.sau-4B laid a foundation for the following fine mapping and cloning work.