Advancing understanding of oat phenology for crop adaptation.

Oat (Avena sativa) is an annual cereal grown for forage, fodder and grain. Seasonal flowering behaviour, or phenology, is a key contributor to the success of oat as a crop. As a species, oat is a vernalization-responsive long-day plant that flowers after winter as days lengthen in spring. Variation in both vernalization and daylength requirements broadens adaptation of oat and has been used to breed modern cultivars with seasonal flowering behaviours suited to different regions, sowing dates and farming practices. This review examines the importance of variation in oat phenology for crop adaptation. Strategies to advance understanding of the genetic basis of oat phenology are then outlined. These include the potential to transfer knowledge from related temperate cereals, particularly wheat (Triticum aestivum) and barley (Hordeum vulgare), to provide insights into the potential molecular basis of variation in oat phenology. Approaches that use emerging genomic resources to directly investigate the molecular basis of oat phenology are also described, including application of high-resolution genome-wide diversity surveys to map genes linked to variation in flowering behaviour. The need to resolve the contribution of individual phenology genes to crop performance by developing oat genetic resources, such as near-isogenic lines, is emphasised. Finally, ways that deeper knowledge of oat phenology can be applied to breed improved varieties and to inform on-farm decision-making are outlined.

Identification of Sclerotinia stem rot resistance quantitative trait loci in a chickpea (Cicer arietinum) recombinant inbred line population.

Sclerotinia stem rot (SSR), caused by Sclerotinia sclerotiorum , is one of the most economically devastating diseases in chickpea (Cicer arietinum L.). No complete resistance is available in chickpea to this disease, and the inheritance of partial resistance is not understood. Two hundred F7 recombinant inbred lines (RILs) derived from a cross between a partially resistant variety PBA HatTrick, and a highly susceptible variety Kyabra were characterised for their responses to SSR inoculation. Quantitative trait locus (QTL) analysis was conducted for the area under the disease progress curve (AUDPC) after RIL infection with S. sclerotiorum . Four QTLs on chromosomes, Ca4 (qSSR4-1, qSSR4-2), Ca6 (qSSR6-1) and Ca7 (qSSR7-1), individually accounted for between 4.2 and 15.8% of the total estimated phenotypic variation for the response to SSR inoculation. Candidate genes located in these QTL regions are predicted to be involved in a wide range of processes, including phenylpropanoid biosynthesis, plant-pathogen interaction, and plant hormone signal transduction. This is the first study investigating the inheritance of resistance to S. sclerotiorum in chickpea. Markers associated with the identified QTLs could be employed for marker-assisted selection in chickpea breeding.

A QTL on the Ca7 chromosome of chickpea affects resistance to the root-lesion nematode Pratylenchus thornei.

The root-lesion nematode Pratylenchus thornei Sher & Allen, 1953 is a damaging parasite of many crop plants, including the grain legume chickpea (Cicer arietinum L.). Within cultivated chickpea, there are no known sources of strong resistance to P. thornei, but some cultivars have partial resistance. In the research reported here, the genetic basis for differences in P. thornei resistance was analysed using a population derived by accelerated single seed descent from a cross between a partially resistant cultivar, PBA HatTrick, and a very susceptible cultivar, Kyabra. A genetic linkage map was constructed from genotyping-by-sequencing data. Two quantitative trait loci were mapped, one on the Ca4 chromosome and one on the Ca7 chromosome. The Ca7 locus had a greater and more consistent effect than the Ca4 locus. Marker assays designed for single nucleotide polymorphisms on Ca7 were applied to a panel of chickpea accessions. Some of these markers should be useful for marker-assisted selection in chickpea breeding. Haplotype analysis confirmed the Iranian landrace ICC14903 to be the source of the resistance allele in PBA HatTrick and indicated that other Australian cultivars inherited the same allele from other Iranian landraces. A candidate region was defined on the Ca7 pseudomolecule. Within that region, 69 genes have been predicted with high confidence. Among these, two have annotations related to biotic stress response. Three others have previously been reported to be expressed in roots of PBA HatTrick and Kyabra, including one that is more highly expressed in PBA HatTrick than in Kyabra. Supplementary Information: The online version contains supplementary material available at 10.1007/s11032-021-01271-8.

Fine mapping of Rha2 in barley reveals candidate genes for resistance against cereal cyst nematode.

KEY MESSAGE: The cereal cyst nematode resistance locus Rha2 was mapped to a 978 kbp region on the long arm of barley chromosome 2H. Three candidate genes are discussed. The cereal cyst nematode (CCN) Heterodera avenae is a soil-borne obligate parasite that can cause severe damage to cereals. This research involved fine mapping of Rha2, a CCN resistance locus on chromosome 2H of barley. Rha2 was previously mapped relative to restriction fragment length polymorphisms (RFLPs) in two mapping populations. Anchoring of flanking RFLP clone sequences to the barley genome assembly defined an interval of 5077 kbp. Genotyping-by-sequencing of resistant and susceptible materials led to the discovery of potentially useful single nucleotide polymorphisms (SNPs). Assays were designed for these SNPs and applied to mapping populations. This narrowed the region of interest to 3966 kbp. Further fine mapping was pursued by crossing and backcrossing the resistant cultivar Sloop SA to its susceptible ancestor Sloop. Evaluation of F2 progeny confirmed that the resistance segregates as a single dominant gene. Genotyping of 9003 BC2F2 progeny identified recombinants. Evaluation of recombinant BC2F3 progeny narrowed the region of interest to 978 kbp. Two of the SNPs within this region proved to be diagnostic of CCN resistance across a wide range of barley germplasm. Fluorescence-based and gel-based assays were developed for these SNPs for use in marker-assisted selection. Within the candidate region of the reference genome, there are nine high-confidence predicted genes. Three of these, one that encodes RAR1 (a cysteine- and histidine-rich domain-containing protein), one that is predicted to encode an acetylglutamate kinase and one that is predicted to encode a tonoplast intrinsic protein, are discussed as candidate genes for CCN resistance.

A locus on barley chromosome 5H affects adult plant resistance to powdery mildew.

Adult plant resistance against plant pathogens is of interest as a means to achieve durable resistance. Prior to this research, the barley lines CLE210 (from Uruguay) and Denar (from the Czech Republic) had been reported to exhibit adult-plant resistance against powdery mildew. Here, populations of doubled haploid lines from crosses of these lines with the susceptible cultivar Baudin were evaluated for powdery mildew resistance in field experiments. Using linkage maps constructed from genotyping-by-sequencing (GBS) data, it was determined that differences in resistance were largely attributable to a region on the long arm of chromosome 5H (5HL). Therefore, KASP™ assays were developed based on GBS tag sequences mapped on that chromosome, providing more reliable genetic maps. In each population, a large-effect QTL was mapped on 5HL. As no sequence variation was detected between CLE210 and Denar in this region of 5HL, the two sources of resistance may be identical by descent in the QTL region and carry the same resistance gene. Marker assays from the QTL region were evaluated on a panel of barley lines, providing information that breeders could use to select assays for use in marker-assisted selection.

Differential expression of the HvCslF6 gene late in grain development may explain quantitative differences in (1,3;1,4)-ß-glucan concentration in barley.

The cellulose synthase-like gene HvCslF6, which is essential for (1,3;1,4)-ß-glucan biosynthesis in barley, collocates with quantitative trait loci (QTL) for grain (1,3;1,4)-ß-glucan concentration in several populations, including CDC Bold × TR251. Here, an alanine-to-threonine substitution (caused by the only non-synonymous difference between the CDC Bold and TR251 HvCslF6 alleles) was mapped to a position within HvCSLF6 that seems unlikely to affect enzyme stability or function. Consistent with this, transient expression of full-length HvCslF6 cDNAs from CDC Bold and TR251 in Nicotianabenthamiana led to accumulation of similar amounts of (1,3;1,4)-ß-glucan accumulation. Monitoring of HvCslF6 transcripts throughout grain development revealed a significant difference late in grain development (more than 30 days after pollination), with TR251 [the parent with higher grain (1,3;1,4)-ß-glucan] exhibiting higher transcript levels than CDC Bold. A similar difference was observed between Beka and Logan, the parents of another population in which a QTL had been mapped in the HvCslF6 region. Sequencing of a putative promoter region of HvCslF6 revealed numerous polymorphisms between CDC Bold and TR251, but none between Beka and Logan. While the results of this work indicate that naturally occurring quantitative differences in (1,3;1,4)-ß-glucan accumulation may be due to cis-regulated differences in HvCslF6 expression, these could not be attributed to any specific DNA sequence polymorphism. Nevertheless, information on HvCslF6 sequence polymorphism was used to develop molecular markers that could be used in barley breeding to select for the desired [low or high (1,3;1,4)-ß-glucan] allele of the QTL.

Poor Homologous Synapsis 1 Interacts with Chromatin but Does Not Colocalise with ASYnapsis 1 during Early Meiosis in Bread Wheat.

Chromosome pairing, synapsis, and DNA recombination are three key processes that occur during early meiosis. A previous study of Poor Homologous Synapsis 1 (PHS1) in maize suggested that PHS1 has a role in coordinating these three processes. Here we report the isolation of wheat (Triticum aestivum) PHS1 (TaPHS1), and its expression profile during and after meiosis. While the TaPHS1 protein has sequence similarity to other plant PHS1/PHS1-like proteins, it also possesses a unique region of oligopeptide repeat units. We show that TaPHS1 interacts with both single- and double-stranded DNA in vitro and provide evidence of the protein region that imparts the DNA-binding ability. Immunolocalisation data from assays conducted using antisera raised against TaPHS1 show that TaPHS1 associates with chromatin during early meiosis, with the signal persisting beyond chromosome synapsis. Furthermore, TaPHS1 does not appear to colocalise with the asynapsis protein (TaASY1) suggesting that these proteins are probably independently coordinated. Significantly, the data from the DNA-binding assays and 3-dimensional immunolocalisation of TaPHS1 during early meiosis indicates that TaPHS1 interacts with DNA, a function not previously observed in either the Arabidopsis or maize PHS1 homologues. As such, these results provide new insight into the function of PHS1 during early meiosis in bread wheat.

The isolation and characterisation of the wheat molecular ZIPper I homologue, TaZYP1.

BACKGROUND: The synaptonemal complex (SC) is a proteinaceous tripartite structure used to hold homologous chromosomes together during the early stages of meiosis. The yeast ZIP1 and its homologues in other species have previously been characterised as the transverse filament protein of the synaptonemal complex. Proper installation of ZYP1 along chromosomes has been shown to be dependent on the axial element-associated protein, ASY1 in Arabidopsis. RESULTS: Here we report the isolation of the wheat (Triticum aestivum) ZYP1 (TaZYP1) and its expression profile (during and post-meiosis) in wild-type, the ph1b deletion mutant as well as in Taasy1 RNAi knock-down mutants. TaZYP1 has a putative DNA-binding S/TPXX motif in its C-terminal region and we provide evidence that TaZYP1 interacts non-preferentially with both single- and double-stranded DNA in vitro. 3-dimensional dual immunofluorescence localisation assays conducted with an antibody raised against TaZYP1 show that TaZYP1 interacts with chromatin during meiosis but does not co-localise to regions of chromatin where TaASY1 is present. The TaZYP1 signal lengthens into regions of chromatin where TaASY1 has been removed in wild-type but this appears delayed in the ph1b mutant. The localisation profile of TaZYP1 in four Taasy1 knock-down mutants is similar to wild-type but TaZYP1 signal intensity appears weaker and more diffused. CONCLUSIONS: In contrast to previous studies performed on plant species where ZYP1 signal is sandwiched by ASY1 signal located on both axial elements of the SC, data from the 3-dimensional dual immunofluorescence localisation assays conducted in this study show that TaZYP1 signal only lengthens into regions of chromatin after TaASY1 signal is being unloaded. However, the observation that TaZYP1 loading appears delayed in both the ph1b and Taasy1 mutants suggests that TaASY1 may still be essential for TaZYP1 to play a role in SC formation during meiosis. These data further suggest that the temporal installation of ZYP1 onto pairing homologous chromosomes in wheat is different to that of other plant species and highlights the need to study this synaptonemal complex protein on a species to species basis.

Preliminary characterisation of two early meiotic wheat proteins after identification through 2D gel electrophoresis proteomics.

Various genetic-based approaches including mutant population screens, microarray analyses, cloning and transgenesis have broadened our knowledge of gene function during meiosis in plants. Nonetheless, these genetic tools are not without inherent limitations. One alternative approach to studying plant meiosis, especially in polyploids such as Triticum aestivum L. (bread wheat), is proteomics. However, protein-based approaches using proteomics have seldom been described, with only two attempts at studying early plant meiosis reported. Here, we report the investigation of early bread wheat meiosis using proteomics. Five differentially expressed protein spots were identified using 2D gel electrophoresis (2DGE) on protein extracts from four pooled stages of meiosis and three genotypes (Chinese Spring wild-type, ph1b and ph2a wheat mutant lines). Tandem mass spectrometry (MS/MS) identification of peptides from these protein spots led to the isolation and characterisation of the full-length clones of a wheat Speckle-type POZ protein, an SF21-like protein and HSP70, and a partial coding sequence of a hexose transporter. Significantly, the putative functions of the Speckle-type POZ protein and HSP70 were confirmed using in vitro DNA binding assays. Through the use of a 2DGE proteomics approach, we show that proteomics is a viable alternative to genetic-based approaches when studying meiosis in wheat. More significantly, we report a potential role for a Speckle-type POZ protein and a HSP70 in chromosome pairing during the early stages of meiosis in bread wheat.

The RAD51 gene family in bread wheat is highly conserved across eukaryotes, with RAD51A upregulated during early meiosis.

The RADiation sensitive protein 51 (RAD51) recombinase is a eukaryotic homologue of the bacterial Recombinase A (RecA). It is required for homologous recombination of DNA during meiosis where it plays a role in processes such as homology searching and strand invasion. RAD51 is well conserved in eukaryotes with as many as four paralogues identified in vertebrates and some higher plants. Here we report the isolation and preliminary characterisation of four RAD51 gene family members in hexaploid (bread) wheat (Triticum aestivum L.). RAD51A1, RAD51A2 and RAD51D were located on chromosome group 7, and RAD51C was on chromosome group 2. Q-PCR gene expression profiling revealed that RAD51A1 was upregulated during meiosis with lower expression levels seen in mitotic tissue, and bioinformatics analysis demonstrated the evolutionary linkages of this gene family to other eukaryotic RAD51 sequences. Western blot analysis of heterologously expressed RAD51 from bread wheat has shown that it is detectable using anti-human RAD51 antibodies and that molecular modelling of the same protein revealed structural conservation when compared with yeast, human, Arabidopsis and maize RAD51A orthologues. This report has widened the knowledge base of this important protein family in plants, and highlighted the high level of structural conservation among RAD51 proteins from various species.