Systemic development of wheat-Thinopyrum elongatum translocation lines and their deployment in wheat breeding for Fusarium head blight resistance.

Fusarium head blight (FHB), mainly caused by Fusarium graminearum, is one of the most destructive diseases of wheat (Triticum aestivum) around the world. FHB causes significant yield losses and reduces grain quality. The lack of resistance resources is a major bottleneck for wheat FHB resistance breeding. As a wheat relative, Thinopyrum elongatum contains many genes that can be used for wheat improvement. Although the novel gene Fhb-7EL was mapped on chromosome 7EL of Th. elongatum, successful transfer of the FHB resistance gene into commercial wheat varieties has not been reported. In this study, we developed 836 wheat-Th. elongatum translocation lines of various types by irradiating the pollen of the wheat-Th. elongatum addition line CS-7EL at the flowering stage, among which 81 were identified as resistant to FHB. By backcrossing the FHB-resistant lines with the main cultivar Jimai 22, three wheat-Th. elongatum translocation lines, Zhongke 1878, Zhongke 166, and Zhongke 545, were successfully applied in wheat breeding without yield penalty. Combining karyotype and phenotype analyses, we mapped the Fhb-7EL gene to the distal end of chromosome 7EL. Five molecular markers linked with the FHB resistance interval were developed, which facilitates molecular marker-assisted breeding. Altogether, we successfully applied alien chromatin with FHB resistance from Th. elongatum in wheat breeding without yield penalty. These newly developed FHB-resistant wheat-Th. elongatum translocation lines, Zhongke 1878, Zhongke 166, and Zhongke 545, can be used as novel resistance resources for wheat breeding.

Introgression of tetraploid Thinopyrum elongatum 6EL segments enhances the stripe rust resistance of adult wheat plants.

Preventing the widespread occurrence of stripe rust in wheat largely depends on the identification of new stripe rust resistance genes and the breeding of cultivars with durable resistance. In previous study, we reported 6E of wheat-tetraploid Thinopyrum elongatum 6E (6D) substitution line contains adult-stage stripe rust resistance genes. In this study, three novel wheat-tetraploid Th. elongatum translocation lines were generated from the offspring of a cross between common wheat and the 6E (6D) substitution line. Genomic in situ hybridization (GISH), fluorescence in situ hybridization chromosome painting (FISH painting), repetitive sequential FISH, and 55 K SNP analyses indicated that K227-48, K242-82, and K246-6 contained 42 chromosomes and were 6DL·6ES, 2DL·6EL, and 6DS·6EL translocation lines, respectively. The assessment of stripe rust resistance revealed that K227-48 was susceptible to a mixture of Pst races, whereas the 6EL lines K242-82 and K246-6 were highly resistance to stripe rust at the adult stage. Thus, this resistance was due to the chromosome arm 6EL of tetraploid Th. elongatum. The improved agronomic performance of 6DS·6EL translocation line may be a useful novel germplasm resource for wheat breeding programs. For the application of marker-assisted selection (MAS), 47 simple sequence repeat (SSR) markers were developed, showing specific amplification on the chromosome 6E using the whole-genome sequence of diploid Th. elongatum. The 6DS·6EL translocation line and SSR markers have the potential to be deploy for future stripe rust resistance wheat breeding program. Supplementary Information: The online version contains supplementary material available at 10.1007/s11032-024-01493-6.

Cytogenetic and Genomic Characterization of a Novel Wheat-Tetraploid Thinopyrum elongatum 1BS⋠1EL Translocation Line with Stripe Rust Resistance.

Stripe rust, caused by Puccinia striiformis f. sp. tritici, is a destructive wheat disease pathogen. Thinopyrum elongatum is a valuable germplasm including diploid, tetraploid, and decaploid with plenty of biotic and abiotic resistance. In a previous study, we generated a stripe rust-resistant wheat-tetraploid Th. elongatum 1E/1D substitution line, K17-841-1. To further apply the wild germplasm for wheat breeding, we selected and obtained a new homozygous wheat-tetraploid Th. elongatum translocation line, T1BS⋅1EL, using genomic in situ hybridization, fluorescence in situ hybridization (FISH), oligo-FISH painting, and the wheat 55K single nucleotide polymorphism genotyping array. The T1BS⋅1EL is highly resistant to stripe rust at the seedling and adult stages. Pedigree and molecular marker analyses revealed that the resistance gene was located on the chromosome arm 1EL of tetraploid Th. elongatum, tentatively named Yr1EL. In addition, we developed and validated 32 simple sequence repeat markers and two kompetitive allele-specific PCR assays that were specific to the tetraploid Th. elongatum chromosome arm 1EL to facilitate marker-assisted selection for alien 1EL stripe rust resistance breeding. This will help us explore and locate the stripe rust resistance gene mapping on the 1E chromosome and deploy it in the wheat breeding program.

Using Transcriptomics to Determine the Mechanism for the Resistance to Fusarium Head Blight of a Wheat-Th. elongatum Translocation Line.

Fusarium head blight (FHB), caused by the Fusarium graminearum species complex, is a destructive disease in wheat worldwide. The lack of FHB-resistant germplasm is a barrier in wheat breeding for resistance to FHB. Thinopyrum elongatum is an important relative that has been successfully used for the genetic improvement of wheat. In this study, a translocation line, YNM158, with the YM158 genetic background carrying a fragment of diploid Th. elongatum 7EL chromosome created using 60Co-γ radiation, showed high resistance to FHB under both field and greenhouse conditions. Transcriptome analysis confirmed that the horizontal transfer gene, encoding glutathione S-transferase (GST), is an important contributor to FHB resistance in the pathogen infection stage, whereas the 7EL chromosome fragment carries other genes regulated by F. graminearum during the colonization stage. Introgression of the 7EL fragment affected the expression of wheat genes that were enriched in resistance pathways, including the phosphatidylinositol signaling system, protein processing in the endoplasmic reticulum, plant-pathogen interaction, and the mitogen-activated protein kinase (MAPK) signaling pathway at different stages after F. graminearium infection. This study provides a novel germplasm for wheat resistance to FHB and new insights into the molecular mechanisms of wheat resistance to FHB.

Long-range assembly of sequences helps to unravel the genome structure and small variation of the wheat-Haynaldia villosa translocated chromosome 6VS.6AL.

Genomics studies in wild species of wheat have been limited due to the lack of references; however, new technologies and bioinformatics tools have much potential to promote genomic research. The wheat-Haynaldia villosa translocation line T6VS·6AL has been widely used as a backbone parent of wheat breeding in China. Therefore, revealing the genome structure of translocation chromosome 6VS·6AL will clarify how this chromosome formed and will help to determine how it affects agronomic traits. In this study, chromosome flow sorting, NGS sequencing and Chicago long-range linkage assembly were innovatively used to produce the assembled sequences of 6VS·6AL, and gene prediction and genome structure characterization at the molecular level were effectively performed. The analysis discovered that the short arm of 6VS·6AL was actually composed of a large distal segment of 6VS, a small proximal segment of 6AS and the centromere of 6A, while the collinear region in 6VS corresponding to 230-260 Mb of 6AS-Ta was deleted when the recombination between 6VS and 6AS occurred. In addition to the molecular mechanism of the increased grain weight and enhanced spike length produced by the translocation chromosome, it may be correlated with missing GW2-V and an evolved NRT-V cluster. Moreover, a fine physical bin map of 6VS was constructed by the high-throughput developed 6VS-specific InDel markers and a series of newly identified small fragment translocation lines involving 6VS. This study will provide essential information for mining of new alien genes carried by the 6VS·6AL translocation chromosome.

Molecular Cytogenetic Analysis of the Introgression between Agropyron cristatum P Genome and Wheat Genome.

Agropyron cristatum (2n = 4x = 28, PPPP) is an important wild relative of common wheat (Triticum aestivum L., 2n = 6x = 42). A previous report showed that the wheat-A. cristatum 6P translocation line WAT655 carrying A. cristatum 6PS (0.81-1.00) exhibited high resistance to prevalent physiological races of stripe rust (CYR32 and CYR33). In this study, three disease resistance-related transcripts, which were mapped to A. cristatum 6PS (0.81-1.00) through the analysis of specific molecular markers, were acquired from among A. cristatum full-length transcripts. The BC5F2 and BC5F2:3 genetic populations of the translocation line WAT655 were analyzed by using three disease resistance-related gene markers, A. cristatum P genome-specific markers, and fluorescence in situ hybridization (FISH). The results revealed that the introgression between A. cristatum P genome and wheat genome was observed in progenies of the genetic populations of the translocation line WAT655 and the physical positions of the three genes were considerably adjacent on A. cristatum 6PS (0.81-1.00) according to the FISH results. Additionally, kompetitive allele-specific PCR (KASP) markers of the three genes were developed to detect and acquire 24 breeding lines selected from the progenies of the distant hybridization of wheat and A. cristatum, which showed resistance to physiological races of stripe rust (CYR32 and CYR33) and other desirable agronomic traits according to the field investigation. In conclusion, this study not only provides new insights into the introgression between A. cristatum P genome and wheat genome but also provides the desirable germplasms for breeding practice.

Molecular cytogenetic characterization of a novel wheat-Psathyrostachys huashanica Keng T3DS-5NsL•5NsS and T5DL-3DS•3DL dual translocation line with powdery mildew resistance.

BACKGROUND: Psathyrostachys huashanica Keng (2n = 2x = 14, NsNs) carries many outstanding agronomic traits, therefore is a valuable resource for wheat genetic improvement. Wheat-P. huashanica translocation lines are important intermediate materials for wheat breeding and studying the functions of alien chromosomes. However, powdery mildew resistance in these translocation lines has not been reported previously. RESULTS: This study developed a novel wheat-P. huashanica translocation line TR77 by selecting a F7 progeny from the cross between heptaploid hybrid H8911 (2n = 7x = 49, AABBDDNs) and durum wheat line Trs-372. Chromosome karyotype of 2n = 42 = 21II was observed in both mitotic and meiotic stages of TR77. Genomic in situ hybridization analysis identified two translocated chromosomes that paired normally at meiosis stage in TR77. Molecular marker analysis showed that part of chromosome 5D was replaced by part of alien chromosome fragment 5Ns. It meant replacement made part 5DL and part 5NsL·5NsS existed in wheat background, and then translocation happened between these chromosomes and wheat 3D chromosome. Fluorescence in situ hybridization demonstrated that TR77 carries dual translocations: T3DS-5NsL·5NsS and T5DL-3DS·3DL. Analysis using a 15 K-wheat-SNP chip confirmed that SNP genotypes on the 5D chromosome of TR77 matched well with these of P. huashanica, but poorly with common wheat line 7182. The translocation was physically located between 202.3 and 213.1 Mb in 5D. TR77 showed longer spikes, more kernels per spike, and much better powdery mildew resistance than its wheat parents: common wheat line 7182 and durum wheat line Trs-372. CONCLUSIONS: TR77 is a novel stable wheat-P. huashanica T3DS-5NsL·5NsS and T5DL-3DS·3DL dual translocation line and showed significant improved spike traits and resistance to powdery mildew compared to its parents, thus, it can be an useful germplasm for breeding disease resistance and studying the genetic mechanism of dual translocations.

Molecular cytogenetic characterization of wheat-Elymus repens chromosomal translocation lines with resistance to Fusarium head blight and stripe rust.

BACKGROUND: Fusarium head blight (FHB) caused by the fungus Fusarium graminearum Schwabe and stripe rust caused by Puccinia striiformis f. sp. tritici are devastating diseases that affect wheat production worldwide. The use of disease-resistant genes and cultivars is the most effective means of reducing fungicide applications to combat these diseases. Elymus repens (2n = 6x = 42, StStStStHH) is a potentially useful germplasm of FHB and stripe rust resistance for wheat improvement. RESULTS: Here, we report the development and characterization of two wheat-E. repens lines derived from the progeny of common wheat-E. repens hybrids. Cytological studies indicated that the mean chromosome configuration of K15-1192-2 and K15-1194-2 at meiosis were 2n = 42 = 0.86 I + 17.46 II (ring) + 3.11 II (rod) and 2n = 42 = 2.45 I + 14.17 II (ring) + 5.50 II (rod) + 0.07 III, respectively. Genomic and fluorescence in situ hybridization karyotyping and simple sequence repeats markers revealed that K15-1192-2 was a wheat-E. repens 3D/?St double terminal chromosomal translocation line. Line K15-1194-2 was identified as harboring a pair of 7DS/?StL Robertsonian translocations and one 3D/?St double terminal translocational chromosome. Further analyses using specific expressed sequence tag-SSR markers confirmed that the wheat-E. repens translocations involved the 3St chromatin in both lines. Furthermore, compared with the wheat parent Chuannong16, K15-1192-2 and K15-1194-2 expressed high levels of resistance to FHB and stripe rust pathogens prevalent in China. CONCLUSIONS: Thus, this study has determined that the chromosome 3St of E. repens harbors gene(s) highly resistant to FHB and stripe rust, and chromatin of 3St introgressed into wheat chromosomes completely presented the resistance, indicating the feasibility of using these translocation lines as novel material for breeding resistant wheat cultivars and alien gene mining.

Deletion mapping and verification of an enhanced-grain number per spike locus from the 6PL chromosome arm of Agropyron cristatum in common wheat.

KEY MESSAGE: An enhanced-grain number per spike locus from Agropyron cristatum 6PL was mapped onto 6PL (0.27-0.51) via deletion mapping, and its effect was further verified by evaluating a newly created translocation line. Agropyron cristatum (2n = 4x = 28, PPPP) is an important wild relative of common wheat and carries many desirable yield-related traits. The wheat-A. cristatum 6P disomic addition line 4844-12 exhibited high grain number per spike (GNS), high spikelet number per spike (SNS), and high kernel number per spikelet (KNS). In this study, five A. cristatum 6P deletion lines, five wheat-A. cristatum 6P translocation lines, and genetic populations of these lines were used to map the enhanced-GNS locus from A. cristatum chromosome 6P, which were genotyped via genomic in situ hybridization, fluorescence in situ hybridization, or molecular markers. According to the evaluation of the agronomic traits in four growing seasons (2014-2015, 2015-2016, 2016-2017, and 2017-2018), we found that the deletion lines and the translocation lines carrying the long arm of A. cristatum chromosome 6P (6PL) exhibited high GNS, SNS, and KNS, and the enhanced-GNS locus was ultimately mapped onto 6PL (0.27-0.51). To verify the localization results, we created a new translocation line WAT650a (T5BL•5BS-6PL) that carried 6PL (0.35-0.42); this line exhibited higher GNS and SNS than the recipient parent Fukuhokomugi (Fukuho). Collectively, the enhanced-GNS locus of A. cristatum 6PL can be important for improving yield traits in common wheat; the translocation lines with the enhanced-GNS locus can serve as novel and valuable germplasm resources for wheat breeding.

Physical Mapping of Stem Rust Resistance Gene Sr52 from Dasypyrum villosum Based on ph1b-Induced Homoeologous Recombination.

Wheat stem rust caused by Puccinia graminis f. sp. tritici (Pgt) had been a devastating foliar disease worldwide during the 20th century. With the emergence of Ug99 races, which are virulent to most stem rust resistance genes deployed in wheat varieties and advanced lines, stem rust has once again become a disease threatening global wheat production. Sr52, derived from Dasypyrum villosum and mapped to the long arm of 6V#3, is one of the few effective genes against Ug99 races. In this study, the wheat-D. villosum Robertsonian translocation T6AS·6V#3L, the only stock carrying Sr52 released to experimental and breeding programs so far, was crossed with a CS ph1b mutant to induce recombinants with shortened 6V#3L chromosome segments locating Sr52. Six independent homozygous recombinants with different segment sizes and breakpoints were developed and characterized using in situ hybridization and molecular markers analyses. Stem rust resistance evaluation showed that only three terminal recombinants (1381, 1380, and 1392) containing 8%, 22%, and 30% of the distal segment of 6V#3L, respectively, were resistant to stem rust. Thus, the gene Sr52 was mapped into 6V#3L bin FL 0.92-1.00. In addition, three molecular markers in the Sr52-located interval of 6V#3L were confirmed to be diagnostic markers for selection of Sr52 introgressed into common wheat. The newly developed small segment translocation lines with Sr52 and the identified molecular markers closely linked to Sr52 will be valuable for wheat disease breeding.

An intercalary translocation from Agropyron cristatum 6P chromosome into common wheat confers enhanced kernel number per spike.

MAIN CONCLUSION: This study explored 6P chromosomal translocations in wheat, and determined the effects of 6P intercalary chromosome segments on kernel number per wheat spike. Exploiting and utilising gene(s) from wild relative species has become an essential strategy for wheat crop improvement. In the translocation line Pubing2978, the intercalary 6P chromosome segment from Agropyron cristatum (L.) Gaertn. (2n = 4x = 28, PPPP) carried valuable multi-kernel gene(s) and was selected from the offspring of the common wheat plant Fukuho and the irradiated wheat-A. cristatum 6P disomic substitution line 4844-8. Genomic in situ hybridisation (GISH), dual-colour fluorescence in situ hybridisation (FISH), and molecular markers were used to detect the small segmental 6P chromosome in the wheat background and its translocation breakpoint. Cytological studies demonstrated that Pubing2978 was a T1AS-6PL-1AS·1AL intercalary translocation with 42 chromosomes. The breakpoint was located near the centromeric region on the wheat chromosome 1AS and was flanked by the markers SSR12 and SSR283 based on an F2 linkage map. The genotypic data, combined with the phenotypic information, implied that A. cristatum 6P chromosomal segment plays an important role in regulating the kernel number per spike (KPS). By comparison, the mean value of KPS in plants with translocations was approximately 10 higher than that in plants without translocations in three segregated populations. Moreover, the improvement in KPS was likely achieved by increasing both the spikelet number per spike (SNS) and the kernel number per spikelet. These excellent agronomic traits laid the foundation for further investigation of valuable genes and make the Pubing2978 line a promising germplasm for wheat breeding.

Molecular cytogenetic characterization of a new wheat-rye 1BL•1RS translocation line expressing superior stripe rust resistance and enhanced grain yield.

MAIN CONCLUSION: A new wheat-rye 1BL•1RS translocation line, with the characteristics of superior stripe rust resistance and high thousand-kernel weight and grain number per spike, was developed and identified from progenies of wheat-rye- Psathyrostachys huashanica trigeneric hybrids. The wheat-rye 1BL•1RS translocation line from Petkus rye has contributed substantially to the world wheat production. However, due to extensive growing of cultivars with disease resistance genes from short arm of rye chromosome 1R and coevolution of pathogen virulence and host resistance, these cultivars successively lost resistance to pathogens. In this study, a new wheat-rye line K13-868, derived from the progenies of wheat-rye-Psathyrostachys huashanica trigeneric hybrids, was identified and analyzed using fluorescence in situ hybridization (FISH), genomic in situ hybridization (GISH), acid polyacrylamide gel electrophoresis (A-PAGE), and molecular markers. Cytological studies indicated that the mean chromosome configuration of K13-868 at meiosis was 2n = 42 = 0.14 I + 18.78 II (ring) + 2.15 II (rod). Sequential FISH and GISH results demonstrated that K13-868 was a compensating wheat-rye 1BL•1RS Robertsonian translocation line. Acid PAGE analysis revealed that clear specific bands of rye 1RS were expressed in K13-868. Simple sequence repeat (SSR) and rye 1RS-specific markers ω-sec-p1/ω-sec-p2 and O-SEC5'-A/O-SEC3'-R suggested that the 1BS arm of wheat had been substituted by the 1RS arm of rye. At the seedling and adult growth stage, compared with its recurrent wheat parent SM51 and six other wheat cultivars containing the 1RS arm in southwestern China, K13-868 showed high levels of resistance to stripe rust (Puccinia striiformis f. sp. tritici, Pst) pathogens prevalent in China, which are virulent to Yr10 and Yr24/Yr26. In addition, K13-868 expresses higher thousand-kernel weight and more grain number per spike than these controls in two growing seasons, suggesting that this line may carry yield-related genes of rye. This translocation line, with significant characteristics of resistance to stripe rust and high thousand-kernel weight and grain number per spike, could be utilized as a valuable germplasm for wheat improvement.

Characterization of wheat - Psathyrostachys huashanica small segment translocation line with enhanced kernels per spike and stripe rust resistance.

Psathyrostachys huashanica Keng (2n = 2x = 14, NsNs), a distant wild relative of common wheat, possesses rich potentially valuable traits, such as disease resistance and more spikelets and kernels per spike, that could be useful for wheat genetic improvement. Development of wheat - P. huashanica translocation lines will facilitate its practical utilization in wheat breeding. In the present study, a wheat - P. huashanica small segmental translocation line, K-13-835-3, was isolated and characterized from the BC1F5 population of a cross between wheat - P. huashanica amphiploid PHW-SA and wheat cultivar CN16. Cytological studies showed that the mean chromosome configuration of K-13-835-3 at meiosis was 2n = 42 = 0.10 I + 19.43 II (ring) + 1.52 II (rod). GISH analyses indicated that chromosome composition of K-13-835-3 included 40 wheat chromosomes and a pair of wheat - P. huashanica translocation chromosomes. FISH results demonstrated that the small segment from an unidentified P. huashanica chromosome was translocated into wheat chromosome arm 5DS, proximal to the centromere region of 5DS. Compared with the cultivar wheat parent CN16, K-13-835-3 was highly resistant to stripe rust pathogens prevalent in China. Furthermore, spikelets and kernels per spike in K-13-835-3 were significantly higher than those of CN16 in two growing seasons. These results suggest that the desirable genes from P. huashanica were successfully transferred into CN16 background. This translocation line could be used as novel germplasm for high-yield and, eventually, resistant cultivar breeding.

Cytological identification of an Aegilops variabilis chromosome carrying stripe rust resistance in wheat.

Aegilops variabilis (UUSvSv), an important sources for wheat improvement, originated from chromosome doubling of a natural hybrid between Ae. umbellulata (UU) with Ae. longissima (SlSl). The Ae. variabilis karyotype was poorly characterized by fluorescent in situ hybridization (FISH). The FISH probe combination of pSc119.2, pTa71 and pTa-713 identified each of the 14 pairs of Ae. variabilis chromosomes. Our FISH ideogram was further used to detect an Ae. variabilis chromosome carrying stripe rust resistance in the background of wheat lines developed from crosses of the stripe rust susceptible bread wheat cultivar Yiyuan 2 with a resistant Ae. variabilis accession. Among the 15 resistant BC1F7 lines, three were 2Sv + 4Sv addition lines (2n = 46) and 12 were 2Sv(2B) or 2Sv(2D) substitution lines that were confirmed with SSR markers. SSR marker gwm148 can be used to trace 2Sv in common wheat background. Chromosome 2Sv probably carries gametocidal(Gc) gene(s) since cytological instability and chromosome structural variations, including non-homologous translocations, were observed in some lines with this chromosome. Due to the effects of photoperiod genes, substitution lines 2Sv(2D) and 2Sv(2B) exhibited late heading with 2Sv(2D) lines being later than 2Sv(2B) lines. 2Sv(2D) substitution lines were also taller and exhibited higher spikelet numbers and longer spikes.

Transferring an Adult-Plant Stripe-Rust Resistance Gene Yr7VS from Chromosome 7V of Dasypyrum villosum (L.) to Bread Wheat.

Stripe rust (Puccinia striiformis West. f.sp. tritici, Pst) is a destructive disease that seriously threatens wheat production globally. Exploring novel resistance genes for use in wheat breeding is an urgent need, as continuous Pst evolution frequently leads to a breakdown of host resistance. Here, we identified a set of wheat-Dasypyrum villosum 01I139 (V#6) disomic introgression lines for the purpose of determining their responses to a mixture of Pst isolates CYR32, CYR33 and CYR34 at both seedling and adult-plant stages. The results showed that all introgression lines exhibited high susceptibility at the seedling stage, with infection-type (IT) scores in the range of 6-8, whereas, for chromosomes 5V#6 and 7V#6, disomic addition lines NAU5V#6-1 and NAU7V#6-1 displayed high resistance at the adult-plant stage, indicating that adult-plant resistance (APR) genes were located on them. Further, in order to transfer the stripe-rust resistance on chromosome 7V#6, four new wheat-D. villosum introgression lines were identified, by the use of molecular cytogenetic approaches, from the self-pollinated seeds of 7D and 7V#6, in double monosomic line NAU7V#6-2. Among them, NAU7V#6-3 and NAU7V#6-4 were t7V#6L and t7V#6S monosomic addition lines, and NAU7V#6-5 and NAU7V#6-6 were homozygous T7DS·7V#6L and T7DL·7V#6S whole-arm translocation lines. Stripe-rust tests and genetic analyses of chromosome 7V#6 introgression lines revealed a dominant APR gene designated as Yr7VS on the chromosome arm 7V#6S. Comparison with the homozygous T7DL·7V#6S translocation line and the recurrent parent NAU0686 showed no significant differences in yield-related traits. Thus, T7DL·7V#6S whole-arm translocation with the APR gene Yr7VS provided a valuable germplasm for breeding for resistance.

Development and cytological characterization of wheat-Thinopyrum intermedium translocation lines with novel stripe rust resistance gene.

Wheat stripe rust is a destructive disease in many cool and temperate regions around the world. Exploiting novel sources of resistance can provide wheat cultivars with robust and durable resistance to stripe rust. The wheat-Thinopyrum intermedium addition line TAI-14 was proven to carry a stripe rust resistance gene (named as YrT14) on the alien Th. intermedium chromosome. In order to transfer the resistance gene to wheat, wheat-Th. intermedium translocation lines were created by irradiating the pollen of the line TAI-14. We totally obtained 153 wheat-Th. intermedium translocation lines, among which the long alien segmental translocation line Zhongke 78 and the intercalary translocation line Zhongke 15 not only showed good integrated agronomic traits but also were identified as highly resistant to stripe rust in both seedling and adult plant stages. The alien chromatin in Zhongke 15 was identified as an insertion into the satellite of chromosome 6B, a type of translocation never reported before in chromosome engineering. By screening Simple Sequence Repeat (SSR) and Expressed Sequence Tag (EST) markers as well as the markers developed from RNA-sequencing (RNA-Seq) data, 14 markers were identified specific for the alien chromosome and a physical map was constructed. Both Zhongke 78 and Zhongke 15 could be used as a novel source of stripe rust resistance for wheat breeding, and the linked marker T14K50 can be used for molecular marker-assisted breeding. Finally, based on the karyotype, reaction to stripe rust, and genome resequencing data of different wheat-Th. intermedium translocation lines, the stripe rust resistance gene YrT14 was located to an 88.1 Mb interval from 636.7 to 724.8 Mb on Th. intermedium chromosome 19 corresponding to 7J or 7Js.

Molecular Cytogenetic Identification of the Wheat-Dasypyrum villosum T3DL·3V#3S Translocation Line with Resistance against Stripe Rust.

The annual species Dasypyrum villosum possesses several potentially valuable genes for the improvement of common wheat. Previously, we identified a new stripe rust-resistant line, the Chinese Spring (CS)-D. villosum 3V#3 (3D) substitution line (named CD-3), and mapped its potential rust resistance gene (designated as YrCD-3) on the 3V#3 chromosome originating from D. villosum. The objective of the present study was to further narrow down the YrCD-3 locus to a physical region and develop wheat-3V#3 introgression lines with strong stripe rust resistance. By treating CD-3 seeds with 60Co γ-irradiation, two CS-3V#3 translocation lines, T3V#3S.3DL and T3DS.3V#3L (termed 22-12 and 24-20, respectively), were identified from the M4 generation through a combination of non-denaturing fluorescence in situ hybridization (ND-FISH) and functional molecular markers. Stripe rust resistance tests showed that the line 22-12 exhibited strong stripe rust resistance similarly to CD-3, whereas 24-20 was susceptible to stripe rust similarly to CS, indicating that YrCD-3 is located on the short arm of 3V#3. The line 22-12 can potentially be used for further wheat improvement. Additionally, to trace 3V#3 in the wheat genetic background, we produced 30 3V#3-specific sequence tag (EST) markers, among which, 11 markers could identify 3V#3S. These markers could be valuable in fine-mapping YrCD-3.

Development and Molecular Cytogenetic Identification of a New Wheat-Psathyrostachys huashanica Keng Translocation Line Resistant to Powdery Mildew.

Psathyrostachys huashanica Keng, a wild relative of common wheat with many desirable traits, is an invaluable source of genetic material for wheat improvement. Few wheat-P. huashanica translocation lines resistant to powdery mildew have been reported. In this study, a wheat-P. huashanica line, E24-3-1-6-2-1, was generated via distant hybridization, ethyl methanesulfonate (EMS) mutagenesis, and backcross breeding. A chromosome karyotype of 2n = 44 was observed at the mitotic stage in E24-3-1-6-2-1. Genomic in situ hybridization (GISH) analysis revealed four translocated chromosomes in E24-3-1-6-2-1, and P. huashanica chromosome-specific marker analysis showed that the alien chromosome fragment was from the P. huashanica 4Ns chromosome. Moreover, fluorescence in situ hybridization (FISH) analysis demonstrated that reciprocal translocation had occurred between the P. huashanica 4Ns chromosome and the wheat 3D chromosome; thus, E24-3-1-6-2-1 carried two translocations: T3DS·3DL-4NsL and T3DL-4NsS. Translocation also occurred between wheat chromosomes 2A and 4A. At the adult stage, E24-3-1-6-2-1 was highly resistant to powdery mildew, caused by prevalent pathotypes in China. Further, the spike length, numbers of fertile spikelets, kernels per spike, thousand-kernel weight, and grain yield of E24-3-1-6-2-1 were significantly higher than those of its wheat parent 7182 and addition line 24-6-3-1. Thus, this translocation line that is highly resistant to powdery mildew and has excellent agronomic traits can be used as a novel promising germplasm for breeding resistant and high-yielding cultivars.

Cytogenetic identification and molecular marker development for the novel stripe rust-resistant wheat-Thinopyrum intermedium translocation line WTT11.

Stripe rust, caused by Puccinia striiformis f. sp. tritici (Pst), is one of the most destructive diseases of wheat (Triticum aestivum L.) worldwide. Xiaoyan 78829, a partial amphidiploid developed by crossing common wheat with Thinopyrum intermedium, is immune to wheat stripe rust. To transfer the resistance gene of this excellent germplasm resource to wheat, the translocation line WTT11 was produced by pollen irradiation and assessed for immunity to stripe rust races CYR32, CYR33 and CYR34. A novel stripe rust-resistance locus derived from Th. intermedium was confirmed by linkage and diagnostic marker analyses. Molecular cytogenetic analyses revealed that WTT11 carries a TTh·2DL translocation. The breakpoint of 1B was located at 95.5 MB, and the alien segments were found to be homoeologous to wheat-group chromosomes 6 and 7 according to a wheat660K single-nucleotide polymorphism (SNP) array analysis. Ten previously developed PCR-based markers were confirmed to rapidly trace the alien segments of WTT11, and 20 kompetitive allele-specific PCR (KASP) markers were developed to enable genotyping of Th. intermedium and common wheat. Evaluation of agronomic traits in two consecutive crop seasons uncovered some favorable agronomic traits in WTT11, such as lower plant height and longer main panicles, that may be applicable to wheat improvement. As a novel genetic resource, the new resistance locus may be useful for wheat disease-resistance breeding. Supplementary Information: The online version contains supplementary material available at 10.1007/s42994-021-00060-3.