Analysis of Chromosome Associations during Early Meiosis in Wheat Lines Carrying Chromosome Introgressions from Agropyron cristatum

Crested wheatgrass (Agropyron cristatum L. Gaertn., genome P), included in the Triticeae tribe (family Poaceae), is one of the most important grasses in temperate regions. It has been valued as a donor of important agronomic traits for wheat improvement, including tolerance to cold, drought, and high salinity, as well as resistance to leaf rust, stripe rust, and powdery mildew. For successful incorporation of beneficial alleles into wheat, it is essential that recombination between wheat and A. cristatum chromosomes occurs. In this work, we analysed chromosome associations during meiosis in wheat lines carrying chromosome introgressions from A. cristatum chromosomes 5P and 6P in the presence and absence of Ph1 locus using fluorescence in situ hybridisation. The results showed that the Ph1 locus does not affect chromosome associations between A. cristatum and wheat chromosomes because there were no interspecific chromosome associations; therefore, no recombination between chromosomes from wheat and Agropyron were observed in the absence of the Ph1 locus. The 5P and 6P A. cristatum chromosomes do not have a suppressor effect on the Ph1 locus. Wheat univalents in metaphase I suggest that Agropyron chromosomes might carry genes having a role in wheat homologous chromosome associations. Putative effect of the Agropyron genes on wheat chromosome associations does not interact with the Ph1 locus.

Molecular Cytogenetic Identification of a Novel Wheat–Thinopyrum ponticum 1JS (1B) Substitution Line Resistant to Powdery Mildew and Leaf Rust

Thinopyrum ponticum (2n = 10x = 70) is a wild relative of wheat with high tolerance to both biotic and abiotic stresses; it has been wildly used in wheat genetic improvement. A disomic substitution line named SN19647 was derived from a cross between Triticum aestivum and the wheat–Th. ponticum partial amphiploid SNTE20 (2n = 8x = 56). It was evaluated for disease resistance and characterized via sequential fluorescence in situ hybridization (FISH)-genomic in situ hybridization (GISH) and molecular markers. The results showed that SN19647 carried resistance to both powdery mildew and leaf rust. It contained 42 chromosomes with a pair of wheat chromosome 1B replaced by a pair of JS chromosomes from Th. ponticum. In addition to chromosomal substitution events, structural variation also occurred on wheat chromosomes 2A, 5A, 6B, and 7B. Based on marker analysis, 19 markers specific to the JS chromosome were obtained, of which seventeen markers belonged to homoeologous group one. These results indicated that SN19647 was a 1JS (1B) substitution line. Compared with the known 1JS (1D) substitution line CH10A5, it was found that 17 markers generated different specific bands to Th. ponticum, confirming the novelty of the 1JS chromosome in SN19647. Therefore, SN19647, resistant to powdery mildew and leaf rust, was a novel 1JS (1B) substitution line that can be used in wheat genetic improvement.

A novel approach to develop wheat chromosome-specific KASP markers for detecting Amblyopyrum muticum segments in doubled haploid introgression lines

Many wild relative species are being used in pre-breeding programmes to increase the genetic diversity of wheat. Genotyping tools such as single nucleotide polymorphism (SNP)-based arrays and molecular markers have been widely used to characterise wheat-wild relative introgression lines. However, due to the polyploid nature of the recipient wheat genome, it is difficult to develop SNP-based KASP markers that are codominant to track the introgressions from the wild species. Previous attempts to develop KASP markers have involved both exome- and PCR-amplicon-based sequencing of the wild species. But chromosome-specific KASPs assays have been hindered by homoeologous SNPs within the wheat genome. This study involved whole genome sequencing of the diploid wheat wild relative Amblyopyrum muticum and development of a SNP discovery pipeline that generated ~38,000 SNPs in single-copy wheat genome sequences. New assays were designed to increase the density of Am. muticum polymorphic KASP markers. With a goal of one marker per 60 Mbp, 335 new KASP assays were validated as functional. Together with assays validated in previous studies, 498 well distributed chromosome-specific markers were used to recharacterize previously genotyped wheat-Am. muticum doubled haploid (DH) introgression lines. The chromosome specific nature of the KASP markers allowed clarification of which wheat chromosomes were involved with recombination events or substituted with Am. muticum chromosomes and the higher density of markers allowed detection of new small introgressions in these DH lines.

Characterisation of a 4A QTL for Metribuzin Resistance in Wheat by Developing Near-Isogenic Lines

Wheat (Triticum aestivum L.) production is constantly affected by weeds in the farming system. Chemical-based weed management is widely practiced; broad-spectrum herbicides such as metribuzin have been successfully used to control weeds in Australia and elsewhere of the world. Breeding metribuzin-resistant wheat through genetic improvement is needed for effective control of weeds. Quantitative trait loci (QTLs) mapping efforts identified a major QTL on wheat chromosome 4A, explaining up to 20% of the phenotypic variance for metribuzin resistance. The quantitative nature of inheritance of this QTL signifies the importance of near-isogenic lines (NILs), which can convert a quantitative trait into a Mendelian factor for better resolution of the QTL. In the current study, NILs were developed using a heterogeneous inbred family method combined with a fast generation-cycling system in a population of Chuan Mai 25 (resistant) and Ritchie (susceptible). Seven pairs of NILs targeting the 4A QTL for metribuzin resistance were confirmed with a molecular marker and phenotyping. The resistant allele from the resistant parent increased metribuzin resistance by 63–85% (average 69%) compared with the susceptible allele from the susceptible parent. Segregation analysis in the NIL pairs for thousand grain weight (TGW) (g), biomass per plant (kg), tillers per plant, plant height (cm), yield per plant, and powdery mildew visual score (0–9) indicated that these traits were linked with metribuzin resistance. Similarly, TGW was observed to co-segregate with metribuzin resistance in most confirmed NILs, signifying that the two traits are controlled by closely linked genes. The most contrasting NILs can be further characterised by transcriptomic and proteomic analyses to identify the candidate genes responsible for metribuzin resistance.

Analysis of Expression and Bioinformatics of Trehalose-6-phosphate Phosphatases (TPP) Gene Family in Wheat Indicates a Role in Plant Development and Stress Response

Background: Trehalose-6-phosphate phosphatases genes (TPPs) are involved in the development and stress response of plants by regulating the biosynthesis of trehalose, though little is currently known about TPPs in common wheat (Triticum aestivum L.).Results: In this study, we performed a genome-wide identification of the TPP gene family in common wheat, and identified a total of 31 TPP genes. These were subdivided into six subfamilies based on the phylogenetic relationships and the conservation of protein in six monocot and eudicot plants. The majority of TPP genes were represented by 2-3 wheat homoalleles (named TaTPPX_ZA, TaTPPX_ZB, or TaTPPX_ZD), where Z is the location on the wheat chromosome of the gene number (X). We also analyzed the chromosomal location, exon-intron structure, orthologous genes, and protein motifs of the TaTPPs. The RNA-seq data was used to perform an expression analysis, which found 26 TaTPP genes to be differentially expressed based on spatial and temporal characteristics, indicating they have varied functions in the growth and development of wheat. Additionally, we assessed how the promoter regulatory elements were organized and used qRT-PCR in the leaves to observe how they were expressed following ABA, salt, low tempreture, and drought stress treatments. All of these genes exhibited differential expression against one or more stress treatments. Furthermore, overexpressing TaTPP11 in Arabidopsis results delayed plant development and enhanced drought tolerance, but not affect seed morphology. Conclusions: TaTPPs could serve important roles in the development and stress response in wheat. These results provide a basis for subsequent research into the function of TaTPPs.

Genome-Wide Investigation of Spliceosomal SM/LSM Genes in Wheat (Triticum aestivum L.) and Its Progenitors

The SSM/SLSM (spliceosomal Smith (SM)/SM-like (LSM)) genes are the central components of the spliceosome in eukaryotes, which play an important role in regulating RNA splicing, participating in diverse biological processes. Although it has been detected in Arabidopsis and rice etc. plants, the members and significance of the SSM/SLSM gene family in wheat are still not reported. In this study, we identified the SSM/SLSM genes in wheat and its progenitors at genome-scale, where 57 SSM/SLSM genes were identified in wheat, together with 41, 17and 19 found in Triticum dicoccoides, Triticum urartu, and Aegilops tauschii. Furthermore, their phylogenetic relationship, gene structures, conserved motifs, and cis-regulatory elements were systematically analyzed. By synteny analysis, good collinearity of SSM/SLSM genes was found among bread wheat and its progenitors’ genomes, and the distribution of SMD2 genes in wheat chromosome 5A, 4B and 4D located in the 4AL-5AL-7BS chromosome model, due to the translocation. Then, the positively selected genes were further investigated based on the non-synonymous to synonymous (dN/dS) analysis of the orthologous pairs. Finally, the expression profiles of the SSM/SLSM genes were detected using RNA-seq datasets, and eight stress-responsive candidate genes were selected to validate their expression through qPCR (real-time quantitative polymerase chain reaction). According to the co-expression network analysis, the correlation between the LSM7-7A gene and related genes was illustrated through Gene Ontology (GO) enrichment analysis. Furthermore, the LSM7-7A gene was related to the Arabidopsis homologous salt tolerance gene RCY1. This investigation systematically identified the complete candidates of SSM/SLSM genes and their characters in wheat and its progenitors, and provided clues to a better understanding of their contribution during the wheat polyploidy process.

Introgression of Thinopyrum elongatum DNA fragments carrying resistance to fusarium head blight into Triticum aestivum cultivar Chinese Spring is associated with alteration of gene expression.

The tall wheatgrass species <i>Thinopyrum elongatum</i> carries on the long arm of its chromosome 7E a locus contributing strongly to resistance to fusarium head blight (FHB), a devastating fungal disease affecting wheat crops in all temperate areas of the world. Introgression of <i>Th. elongatum</i> 7E chromatin into chromosome 7D of wheat was induced by the <i>ph1b</i> mutant of CS. Recombinants between chromosome 7E and wheat chromosome 7D, induced by the <i>ph1b</i> mutation, were monitored by a combination of molecular markers and phenotyping for FHB resistance. Progeny of up to five subsequent generations derived from two lineages, 64-8 and 32-5, were phenotyped for FHB symptoms and genotyped using published and novel 7D- and 7E-specific markers. Fragments from the distal end of 7EL, still carrying FHB resistance and estimated to be less than 114 and 66 Mbp, were identified as introgressed into wheat chromosome arm 7DL of progeny derived from 64-8 and 32-5, respectively. Gene expression analysis revealed variation in the level of expression of genes from the distal ends of 7EL and 7DL in the introgressed progeny. The 7EL introgressed material will facilitate use of the 7EL FHB resistance locus in wheat breeding programs.