Molecular analyses in wheat and Aegilops tauschii reveals a new orthologue of the leaf rust resistance gene Lr19 on 7DL chromosome of Ae. tauschii

Leaf rust is one of the most devastating wheat diseases worldwide, to which many resistance genes have been ‎successfully introgressed ‎from wheat wild relatives. Though the Thinopyrum ‎ponticum-derived leaf rust resistance gene Lr19, is widely effective worldwide and previous studies ‎have shown its likely presence in Aegilops tauschii, no ‎thorough investigation has been conducted to confirm this. The present study aimed to ‎examine the presence of Lr19 in Ae. tauschii using a collection of molecular and bioinformatic analysis. Accordingly, the Thatcher line was used as susceptible, and a Thatcher+Lr19 (TcLr19) and Agatha were used as resistant lines. CDHLQ pathotyping coupled with DNA markers genotyping verified the presence of an Lr19 orthologue on Ae. tauschii 7DL (AtLr19). Sequencing of the GB marker products from Ae. tauschii and TcLr19 showed 99% homology in these fragments, confirming phenotyping and genotyping results. Both isolated segments were matched to a putative melatonin biosynthesis gene, namely O-methyltransferase-2 (OMT2) mapped to 7DL, with 100% identity. A hierarchical gene network was reconstructed using all identified putative genes within a genomic region containing 2.5 cM upstream and downstream of the OMT2 gene. Results indicated that several numbers of important biotic stress-responsive genes such as RPM1, RGA2, TRIUR3, BURP12, and myosin-11, were located downstream of melatonin as a master regulator molecule through the OMT2 node. To our knowledge, this is the first report of finding an orthologue for ‎Lr19 in Ae. tauschii, which provides insights into the possible regulatory route of LR19.

The barley leaf rust resistance gene Rph3 encodes a putative executor protein

Host resistance is considered the most effective means to control plant diseases; however, individually deployed resistance genes are often rapidly overcome by pathogen adaptation. Combining multiple effective resistance genes is the optimal approach to durable resistance, but the lack of functional markers for resistance genes has hampered implementation. Leaf rust, caused by Puccinia hordei, is an economically significant disease of barley, but only a few major Resistance genes to P. hordei (Rph) have been cloned. In this study, gene Rph3 was isolated by positional cloning and confirmed by mutational analysis and transgenic complementation. The Rph3 gene, which originated from wild barley and was first introgressed into cultivated Egyptian germplasm, encodes a unique transmembrane resistance protein that differs from all known plant disease resistance proteins at the amino acid sequence level. Genetic profiles of diverse accessions indicated limited genetic diversity in Rph3 in domesticated germplasm, and higher diversity in wild barley from the Eastern Mediterranean region. Expression profiling using P. hordei isolates with contrasting pathogenicity for the Rph3 host locus showed that the Rph3 gene was expressed only in interactions with Rph3-avirulent isolates, a phenomenon also observed for transcription activator-like effector-dependent genes known as executors conferring resistance to Xanthomonas spp. Like the known transmembrane executors such as Bs3 and Xa7 heterologous expression of Rph3 in N. benthamiana induced a cell death response. Given that Rph3 shares several features with executor genes, it seems likely that P. hordei contains effectors similar to the transcription activator-like effectors that target host executor genes. The isolation of Rph3 highlights convergent evolutionary processes in diverse plant-pathogen interaction systems, where similar defence mechanisms evolved independently in monocots and dicots and provide evidence for executor genes in the Triticeae tribe.

Fine Mapping of the Leaf Rust Resistance Gene Lr65 in Spelt Wheat ‘Altgold’

Wheat leaf rust (also known as brown rust), caused by the fungal pathogen Puccinia triticina Erikss. (Pt), is one by far the most troublesome wheat disease worldwide. The exploitation of resistance genes has long been considered as the most effective and sustainable method to control leaf rust in wheat production. Previously the leaf rust resistance gene Lr65 has been mapped to the distal end of chromosome arm 2AS linked to molecular marker Xbarc212. In this study, Lr65 was delimited to a 0.8 cM interval between flanking markers Alt-64 and AltID-11, by employing two larger segregating populations obtained from crosses of the resistant parent Altgold Rotkorn (ARK) with the susceptible parents Xuezao and Chinese Spring (CS), respectively. 24 individuals from 622 F2 plants of crosses between ARK and CS were obtained that showed the recombination between Lr65 gene and the flanking markers Alt-64 and AltID-11. With the aid of the CS reference genome sequence (IWGSC RefSeq v1.0), one SSR marker was developed between the interval matched to the Lr65-flanking marker and a high-resolution genetic linkage map was constructed. The Lr65 was finally located to a region corresponding to 60.11 Kb of the CS reference genome. The high-resolution genetic linkage map founded a solid foundation for the map-based cloning of Lr65 and the co-segregating marker will facilitate the marker-assisted selection (MAS) of the target gene.

A membrane-bound ankyrin repeat protein confers race-specific leaf rust disease resistance in wheat

Plasma membrane-associated and intracellular proteins and protein complexes play a pivotal role in pathogen recognition and disease resistance signaling in plants and animals. The two predominant protein families perceiving plant pathogens are receptor-like kinases and nucleotide binding-leucine-rich repeat receptors (NLR), which often confer race-specific resistance. Leaf rust is one of the most prevalent and most devastating wheat diseases. Here, we clone the race-specific leaf rust resistance gene Lr14a from hexaploid wheat. The cloning of Lr14a is aided by the recently published genome assembly of ArinaLrFor, an Lr14a-containing wheat line. Lr14a encodes a membrane-localized protein containing twelve ankyrin (ANK) repeats and structural similarities to Ca2+-permeable non-selective cation channels. Transcriptome analyses reveal an induction of genes associated with calcium ion binding in the presence of Lr14a. Haplotype analyses indicate that Lr14a-containing chromosome segments were introgressed multiple times into the bread wheat gene pool, but we find no variation in the Lr14a coding sequence itself. Our work demonstrates the involvement of an ANK-transmembrane (TM)-like type of gene family in race-specific disease resistance in wheat. This forms the basis to explore ANK-TM-like genes in disease resistance breeding.

Molecular mapping of a new recessive wheat leaf rust resistance gene originating from Triticum spelta

TSD276-2, a wheat genetic stock derived from the cross Agra Local/T. spelta 276 showed broad spectrum resistance against leaf rust pathogen. Genetic analysis was undertaken using F1, F2, F2:3 and BC1F1 generations derived from the cross TSD276-2/Agra Local. The results revealed a single recessive gene for leaf rust resistance, tentatively named as LrTs276-2, in TSD276-2. Molecular mapping of leaf rust resistance gene LrTs276-2 in TSD276-2 was done using SNP-based PCR and SSR markers. For Bulked Segregant Analysis (BSA), two bulks viz. resistant bulk and susceptible bulk, and the parents TSD276-2 and Agra Local were genotyped for SNPs using AFFYMETRIX 35K Wheat Breeders' AXIOM array. T. spelta 276 was also genotyped and used as a check. BSA indicated that the gene for leaf rust resistance in TSD276-2 is located on chromosome arm 1DS. Putatively linked SNPs on chromosome arm 1DS were converted into PCR-based markers. Polymorphic SSR markers on chromosome arm 1DS were also identified. Final linkage map was constructed using one SNP-based PCR and three SSR markers. The rust reaction and chromosomal location suggest that LrTs276-2 is a new leaf rust resistance gene which may be useful in broadening the genetic base of leaf rust resistance in wheat.

Ecological genomics of Chinese wheat improvement: implications in breeding for adaptation

Background China has diverse wheat varieties that adapt to very different environments divided into ten agro-ecological zones. A better understanding of genomic differences and patterns of selection among agro-ecological zones could provide useful information in selection of specific adaptive traits in breeding. Results We genotyped 438 wheat accessions from ten zones with kompetitive allele specific PCR (KASP) markers specific to 47 cloned genes for grain yield, quality, adaptation and stress resistance. Phylogenetic trees and principle component analysis revealed clear differences in winter and spring growth habits. Nucleotide diversity (π) and π ratio (πCL/πMCC) suggested that genetic diversity had increased during breeding, and that Chinese landraces (CL) from Zones I-V contributed little to modern Chinese cultivars (MCC). π ratio and Fst identified 24 KASP markers with 53 strong selection signals specific to Zones I (9 signals), II (12), III (5), IV (5), V (6), and VI (6). Genes with clear genetic differentiation and strong response to selection in at least three zones were leaf rust resistance gene Lr34 (I, II, III and IV), photoperiod sensitivity gene Ppd-D1 (I, II, III, IV and V), vernalization gene Vrn-B1 (V, VII, VIII and X), quality-related gene Glu-B1 (I, II and III) and yield-related genes Sus1-7B (I, II, III, IV and IX), Sus2-2A (I, II, III., IV and VI) and GW2-6B (II, V and VI). Conclusions This study examined selection of multiple genes in each zone, traced the distribution of important genetic variations and provided useful information for ecological genomics and enlightening future breeding goals for different agro-ecological zones.

Characterization of an incomplete leaf rust resistance gene on chromosome 1RS and development of KASP markers for Lr47 in wheat

Leaf rust, caused by Puccininia triticina (Pt), is one of the most common wheat diseases in the Great Plains of the USA. A population of recombinant inbred lines (RILs) from CI 17884 x Bainong 418 was evaluated for responses to leaf rust race Pt52-2 and genotyped using single nucleotide polymorphism (SNP) markers. Quantitative trait locus (QTL) analysis identified a minor gene for resistance to leaf rust, designated QLr.stars-1RS, on the 1BL.1RS translocation segment in Bainong 418, and another leaf rust resistance gene, Lr47, on chromosome 7A of CI 17884. Lr47, originally identified in CI 17884 and located in a wheat-T. speltoides translocation segment 7S#1S, remains one of only a few race-specific resistance genes still effective in the Great Plains. A set of 7A-specific simple sequence repeat (SSR) markers were developed and used to genotype CI 17884 and a pair of near-isogenic lines differing in the presence or absence of 7S#1S, PI 603918 and Pavon F76. Haplotype analysis indicated that the estimated length of 7S#1S was 157.23 to 174.42 Mb, accounting for about 23% of the 7A chromosome. Two SNPs on 7S#1S and 4 SNPs on the 1RS chromosome arm were converted to KASP markers, which were subsequently validated in a panel of cultivars and recently released elite breeding lines. Of these, one and two KASP markers are specific to the 1RS chromosome arm and 7S#1S, respectively, indicating that they can facilitate the introgression of Lr47 and QLr.stars-1BS into locally adapted wheat cultivars and breeding lines.