A cytoplasmic male sterility (CMS) system in durum wheat

Cytoplasmic male sterility (CMS) systems are based in the incompatible interaction between nucleus and cytoplasm and are commonly used for hybrid seed production in many crop species. The msH1 CMS system in common wheat results from the incompatibility between the nuclear genome of wheat and the cytoplasm of Hordeum chilense. Fertility restoration of the CMS phenotype is associated with the addition of the short arm of chromosome 6H^ch from H. chilense. In this work, we attempt to transfer the msH1 system to durum wheat and to evaluate its potential as a new source of CMS for the production of hybrid durum wheat. For that purpose, an alloplasmic durum wheat line was developed by substituting wheat cytoplasm by that from H. chilense. This line was completely male sterile. Also, the double translocation T6H^chS·6DL was transferred from common wheat into durum wheat, to test its potential as a restorer line. Finally, the system was tested by using the double T6H^chS·6DL translocation in durum wheat as pollen donor for the alloplasmic male sterile line, which confirmed the fertility restoration ability of 6H^chS in durum wheat.     

Molecular and cytological characterization of an extra acrocentric chromosome that restores male fertility of wheat in the msH1 CMS system

A new CMS system designated as ‘msH1’ has been reported in bread wheat using the cytoplasm of H. chilense. While testing this system in different wheat backgrounds, a highly fertile line with chromosome number 42 plus an extra acrocentric chromosome was obtained. The extra chromosome did not pair with any wheat chromosome at meiosis, and progeny from this line which lack the acrocentric chromosome showed pollen abortion and male sterility. In order to establish the origin of this chromosome, FISH using H. chilense genomic DNA as probe was used and showed that it had originated from H. chilense chromosome(s). The novel chromosome did not possess sequences similar to wheat rDNA; however, the probe pSc119.2 from S. cereale containing the 120 bp family was found to occur at the end of its long arm. Data obtained from FISH and EST molecular markers confirm that the long arm of the acrocentric chromosome is indeed, the short arm of chromosome 1H^ch from H. chilense. We suggest that the novel chromosome originated from a deletion of the distal part of the long arm of chromosome 1H^ch. Neither the 1H^chS short arm, nor the whole chromosome 1H^ch restores pollen fertility of the alloplasmic wheat. Therefore, the restorer gene on the acrocentric chromosome must be located on the retained segment from the hypothetical 1H^chL, while some pollen fertility inhibitor could be present on the deleted 1H^chL distal segment. Disomic addition of the acrocentric chromosome was obtained and this line resulted fully stable and fertile.     

Novel Bread Wheat Lines Enriched in Carotenoids Carrying Hordeum chilense Chromosome Arms in the ph1b Background

The use of crop wild relative species to improve major crops performance is well established. Hordeum chilense has a high potential as a genetic donor to increase the carotenoid content of wheat. Crosses between the 7H^ch H. chilense substitution lines in wheat and the wheat pairing homoeologous1b (ph1b) mutant allowed the development of wheat-H. chilense translocation lines for both 7H^chα and 7H^chβ chromosome arms in the wheat background. These translocation lines were characterized by in situ hybridization and using molecular markers. In addition, reverse phase chromatography (HPLC) analysis was carried out to evaluate the carotenoid content and both 7H^chα∙7AL and 7AS∙7H^chβ disomic translocation lines. The carotenoid content in 7H^chα∙7AL and 7AS∙7H^chβ disomic translocation lines was higher than the wheat-7H^ch addition line and double amount of carotenoids than the wheat itself. A proteomic analysis confirmed that the presence of chromosome 7H^ch introgressions in wheat scarcely altered the proteomic profile of the wheat flour. The Psy1 (Phytoene Synthase1) gene, which is the first committed step in the carotenoid biosynthetic pathway, was also cytogenetically mapped on the 7H^chα chromosome arm. These new wheat-H. chilense translocation lines can be used as a powerful tool in wheat breeding programs to enrich the diet in bioactive compounds.     

Chromosomal location of structural genes controlling isozymes in Hordeum chilense

Summary Polyacrylamide and starch gel electrophoresis of esterase (EST), glutamate oxaloacetate transaminase (GOT) and phosphoglucomutase (PGM) isozymes in Hordeum chilense, Triticum turgidum conv. durum, the amphiploid H. chilense X T. turgidum (Tritordeum), and the durum wheat/H. chilense monosomic addition lines revealed the chromosomal location of one EST locus, two GOT loci and one PGM locus. Loci Est-H ^ch1 and Got-H ^ch2 were found on chromosome 6H^ch,Got-H ^ch3 on chromosome 3H^ch, and Pgm-H ^ch1 on chromosome 4H^ch. These results lend evidence for the assumed homoeology relationships between chromosomes of Triticeae species.     

Chromosomal location of structural genes controlling isozymes in Hordeum chilense

Summary A study of 6-phosphogluconate dehydrogenase and malate dehydrogenase isozyme expression in Triticum turgidum conv. durum /Hordeum chilense monosomic addition lines has revealed the location of two structural genes, 6-pgd-H ^ch 2 and Mdh-H ^ch 1, on chromosome 1H^ch of H. chilense. The homoeology between 1H^ch and other chromosome of Triticeae related species is discussed on the basis of isozyme gene analysis.     

The genetic control of grain esterases in hexaploid wheat

Summary A comparison of EST-5 grain esterase phenotypes from wheat-alien amphiploid, addition and substitution genotypes, resolved by flat-bed isoelectric focusing identified homoeologous Est-5 loci on chromosome 3H of Hordeum vulgare, 3H^ch of H. chilense, 3S^b of Aegilops bicornis, 3S¹ of Ae. sharonensis and Ae. longissima and 6R of Secale cereale and 6R^m of S. montanum. The Est-5 genes in alien species provide evidence for chromosome homoeology with wheat.     

IRAP,REMAP and ISSR Fingerprinting in Newly Formed Hexaploid Tritordeum (X Tritordeum Ascherson et Graebner) and Respective Parental Species

Retrotransposon (RTN)-based markers, such as the inter-retrotransposon amplified polymorphism (IRAP) and the retrotransposon-microsatellite amplified polymorphism (REMAP), are highly informative, multilocus, and reveal insertion polymorphisms among individuals. These markers have been used for evolutionary studies, genetic diversity assessment, DNA fingerprinting, and detection of genetic rearrangements induced by allopolyploidization. The hexaploid tritordeum (H^chH^chAABB; 2n?=?6x?=?42) is an allopolyploid produced from crosses between wild barley (Hordeum chilense Roem. et Schultz.) (H^chH^ch; 2n?=?2x?=?14) and durum wheat (Triticum turgidum L. conv. durum) (AABB; 2n?=?4x?=?28). With this study, we carried out the DNA fingerprinting of two newly formed hexaploid tritordeum lines (HT22 and HT27) and their respective parents, line H1 of H. chilense and line T81 of durum wheat, based on IRAPs, REMAPs and inter-simple sequence repeats (ISSRs), in order to detect potential rearrangements in tritordeum derived from polyploidization. The amphiploid nature of the HT22 and HT27 individuals was successfully confirmed after fluorescence in situ hybridization (FISH), which was performed on their mitotic chromosome spreads with genomic DNA from H. chilense and 45S ribosomal DNA (rDNA), simultaneously, as probes. Six combinations of LTR (long terminal repeat) primers and seven combinations of one LTR and one SSR (simple sequence repeat) primers successfully produced IRAPs and REMAPs, respectively, in both tritordeum lines, and their respective parents. ISSRs were produced with three SSR primers (8081, 8082, and 8564). The analysis of the presence/absence of bands among the tritordeum lines and the respective parents allowed the detection of polymorphic bands: (1) shared by tritordeum and one of the parents; (2) exclusively amplified in tritordeum; and (3) exclusively present in one of the parents. Once no polymorphism was detected among the individuals of each parental species, the polymorphic bands that fit into the second and third cases probably constituted rearrangements in the newly formed tritordeums that arose in response to allopolyploidization, which resulted from the loss of parental bands or, conversely, from the appearance of novel bands not seen in the parental species. Most of the polymorphic IRAPs in tritordeum were shared with the female parent (H. chilense), while most of the polymorphic REMAPs and ISSRs were common to the male parent (durum wheat), but globally, most of the bands inherited by tritordeum had a wheat origin. In conclusion, these dominant markers were successful for DNA fingerprinting and detection of rearrangements in newly formed tritordeum derived from responses to allopolyploidization.     

Molecular cytogenetic characterization of alien introgressions with gene Fhb3 for resistance to Fusarium head blight disease of wheat

Fusarium head blight (FHB) resistance was identified in the alien species Leymus racemosus, and wheat-Leymus introgression lines with FHB resistance were reported previously. Detailed molecular cytogenetic analysis of alien introgressions T01, T09, and T14 and the mapping of Fhb3, a new gene for FHB resistance, are reported here. The introgression line T09 had an unknown wheat-Leymus translocation chromosome. A total of 36 RFLP markers selected from the seven homoeologous groups of wheat were used to characterize T09 and determine the homoeologous relationship of the introgressed Leymus chromosome with wheat. Only short arm markers for group 7 detected Leymus-specific fragments in T09, whereas 7AS-specific RFLP fragments were missing. C-banding and genomic in situ hybridization results indicated that T09 has a compensating Robertsonian translocation T7AL·7Lr#1S involving the long arm of wheat chromosome 7A and the short arm of Leymus chromosome 7Lr#1 substituting for chromosome arm 7AS of wheat. Introgression lines T01 (2n = 44) and T14 (2n = 44) each had two pairs of independent translocation chromosomes. T01 had T4BS·4BL-7Lr#1S + T4BL-7Lr#1S·5Lr#1S. T14 had T6BS·6BL-7Lr#1S + T6BL·5Lr#1S. These translocations were recovered in the progeny of the irradiated line Lr#1 (T5Lr#1S·7Lr#1S). The three translocation lines, T01, T09, and T14, and the disomic addition 7Lr#1 were consistently resistant to FHB in greenhouse point-inoculation experiments, whereas the disomic addition 5Lr#1 was susceptible. The data indicated that at least one novel FHB resistance gene from Leymus, designated Fhb3, resides in the distal region of the short arm of chromosome 7Lr#1, because the resistant translocation lines share a common distal segment of 7Lr#1S. Three PCR-based markers, BE586744-STS, BE404728-STS, and BE586111-STS, specific for 7Lr#1S were developed to expedite marker-assisted selection in breeding programs.     

OPTIMAS-DW: A comprehensive transcriptomics,metabolomics, ionomics,proteomics and phenomics data resource for maize

Background

The wild barley Hordeum chilense fulfills some requirements for being a useful tool to investigate the endosperm yellow pigment content (YPC) in the Triticeae including its diploid constitution, the availability of genetic resources (addition and deletion stocks and a high density genetic map) and, especially, its high seed YPC not silenced in tritordeums (amphiploids derived from H. chilense and wheat). Thus, the aim of this work was to test the utility of the H. chilense genome for investigating the YPC in the Triticeae.

Results

Twelve genes related to endosperm carotenoid content and/or YPC in grasses (Dxr, Hdr [synonym ispH], Ggpps1, Psy2, Psy3, Pds, Zds, e-Lcy, b-Lcy, Hyd3, Ccd1 and Ppo1) were identified, and mapped in H. chilense using rice genes to identify orthologs from barley, wheat, sorghum and maize. Macrocolinearity studies revealed that gene positions were in agreement in H. vulgare and H. chilense. Additionally, three main regions associated with YPC were identified in chromosomes 2H^ch, 3H^ch and 7H^ch in H. chilense, the former being the most significant one.

Conclusions

The results obtained are consistent with previous findings in wheat and suggest that Ggpps1, Zds and Hyd3 on chromosome 2H^ch may be considered candidate genes in wheat for further studies in YPC improvement. Considering the syntenic location of carotenoid genes in H. chilense, we have concluded that the H^ch genome may constitute a valuable tool for YPC studies in the Triticeae.

     

Allelic variation of the D-prolamin subunits encoded at the Hch genome in a collection of primary hexaploid tritordeums

 Hexaploid tritordeum is the amphiploid derived from the cross between Hordeum chilense and durum wheat. The storage proteins synthesised by the H^ch genome have an influence on the gluten strength of this amphiploid. The D-prolamins of H. chilense are glutenin-like proteins. The variability has been analysed electrophoretically and up to 20 different patterns have been detected in a world collection of this species. This genetic variability of H. chilense could be a source of additional variation in tritordeum and in breeding wheat for quality. Received: 29 November 1998 / Accepted: 19 December 1998     

Linkage relationships between prolamin genes located on chromosome 1H<Superscript>ch</Superscript> in<Emphasis Type="Italic"> Hordeum chilense</Emphasis>

The endosperm storage proteins of Hordeum chilense Roem. et Schult., a species used in the synthesis of the amphiploid tritordeum (×Tritordeum Ascherson et Graebner), have a great effect on the gluten strength of this amphiploid. We have analysed electrophoretically the heredity of these proteins, which are synthesised by genes located on chromosome 1H^ch, and detected up to five loci in a cross between two lines of H. chilense. These loci present a certain homology with loci synthesising the same proteins in wheat. The genetic distances between these loci were calculated.Communicated by H.F. Linskens     

Cereal cyst nematode resistance gene <Emphasis Type="Italic">CreV</Emphasis> effective against <Emphasis Type="Italic">Heterodera filipjevi</Emphasis> transferred from chromosome 6VL of <Emphasis Type="Italic">Dasypyrum villosum</Emphasis> to bread wheat

Cereal cyst nematodes (CCN) are a global economic problem for cereal production. Heterodera filipjevi is one of the most commonly identified and widespread CCN species found in many wheat production regions of the world. Transferring novel genes for resistance to H. filipjevi from wild relatives of wheat is a promising strategy for protection of wheat crops. A set of wheat–Dasypyrum villosum chromosome addition lines, T6V#4S·6AL translocation lines and their donor parental lines were tested for their response to the nematode. D. villosum and wheat–D. villosum disomic addition line DA6V#4 were resistant. As T6V#4S·6AL translocation lines were susceptible, resistance was presumed to be located on chromosome 6V#4L. The objective of this study was to produce and characterize wheat–6V#4L translocations and confirm the chromosome location of the resistance. Introgression lines T6V#4L·6AS, T6V#4L-4BL·4BS and DT6V#4L were developed and subjected to molecular cytogenetic analysis. These and four additional wheat–6V#4 introgression lines were tested for response to H. filipjevi in the greenhouse. The results indicated that introgression lines DA6V#4, T6V#4L·6AS, T6V#4L-4BL·4BS, T6V#4L·6V#4S-7BS and DT6VL#4 had higher levels of H. filipjevi resistance than their recurrent parent. However, Del6V#4L-1 and translocation line T6V#4S·6AL were equally susceptible to wheat cv. Chinese Spring. The CCN resistance gene, temporarily named CreV, was therefore physically mapped to chromosome arm 6V#4L FL 0.80–1.00. Translocation chromosomes T6V#4L·6AS transferred to a modern wheat cv. Aikang 58 with its co-dominant molecular markers could be utilized as a novel germplasm for CCN resistance breeding in wheat.     

Characteristics of alpha-amylase isozymes in cytologenetically different wheat cultivars

The isoenzyme composition of alpha-amylase is studied by polyacrylamide gel electrophoresis in Tris-glycine (pH 8.3) system in wheat cultivars with different genome composition. We show that durum wheat (Triticum durum, 2n = 4x = 28, BBAA) lacks the isoenzymes encoded by 6D and 7D chromosomes that are present in common wheat zymograms (Triticum aestivum, 2n = 6x = 42, BBAADD). A similar pattern is observed in a synthetic allohexaploid carrying the BBAA genomes of wheat and the H ^ch H ^ch genome of barley (Hordeum chilense). Our method of electrophoresis fails to reveal additional variants of alpha-amylase encoded by the barley genome, although C-banding analysis confirms the genomic structure BBAAH ^ch H ^ch of this allopolyploid. The electrophoretic spectrum of the spring common wheat cultivar Dobrynya with the wheat-Agropyron translocation 7DL-7AiL contains all of the alpha-amylase isoenzymes typical for common wheat (2n = 6x = 42, BBAADD) except for the zymotype encoded by the long arm of chromosome 7D. This observation confirms the results of cytogenetic analysis that identified a 7DL-7AiL translocation in this cultivar. No additional alpha-amylase isoenzymes encoded by Agropyron chromosome have been observed. Our data indicate that analysis of wheat-alien hybrids or introgressive forms should be carried out using a complex of different methods.     

Subunits of tetrameric α-amylase inhibitors of Hordeum chilense are encoded by genes located in chromosomes 4Hch and 7Hch

Summary Three proteins (components 1, 2, and 4) of the non-prolamin, 70% ethanol soluble fraction from the endosperm of Hordeum chilense have been identified as putative subunits of the tetrameric inhibitors active against insect -amylases. In experiments carried out with the synthetic alloploid Tritordeum (H. chilense x Triticum turgidum conv. durum), previously described proteins from T. turgidum, designated CM2, CM3 and CM 16, have been also identified as subunits of -amylase inhibitors. Genes for components 1 and 4 of H. chilense have been located in chromosomes 4H^ch and 7H^ch, based on the analysis of H. chilense-T.turgidum addition lines. Subunits of the inhibitors from wheat and from cultivated barley had been previously assigned to chromosomes of the same homoeology groups.     

Chromosomal location of genes coding for endosperm proteins of Hordeum chilense,determined by two-dimensional electrophoresis of wheat-H. chilense chromosome addition lines

The proteins of Hordeum chilense grain were resolved into 25 major components by two-dimensional electrophoresis. Their solubilities in aqueous alcohol solutions were determined to distinguish prolamin storage proteins from metabolic and structural proteins. The prolamins were divided into two groups, based on the presence or absence of intermolecular disulfide bonds determined by gel-filtration chromatography. Using an incomplete set of Chinese Spring wheat-H. chilense disomic addition lines, the structural genes of 21 of the 26 most dominant seed proteins were assigned to chromosomes. The great majority of the prolamin genes, including those coding for a high molecular weight (HMW) prolamin subunit, was present on chromosome 1H^ch. However, a small number of prolamin genes also occurred on chromosomes 5H^ch and 7H^ch. A minor protein, probably belonging to the nonstorage group of proteins, is coded by genes on 5H^ch. Various ditelosomic addition lines and ditelosomic and disomic substitution lines for chromosome 7H^ch were also analyzed by electrophoresis. This technique revealed that the genes for three major prolamins occur on the arm of chromosome 7H^ch and that a gene for a minor protein, also thought to be a prolamin, occurs on the arm. These results are discussed in relation to the evolution of prolamin genes in the Triticeae.     

Meiotic pairing of the amphiploid Hordeum chilense X Triticum turgidum conv. durum studied by means of Giemsa C-banding technique

Summary The meiotic behaviour of the amphiploid Hordeum chilense X Triticum turgidum conv. durum using a C-banding staining method is studied. Nine pairs of chromosomes at metaphase-1 (4A, 7A and the seven of the B genome) were identified and the remaining wheat chromosomes (1A, 2A, 3A, 5A and 6A) and seven of the chilense (1 to 7 H ^ch chromosomes) were assigned to its particular genome. A similar mean number of univalents from parental genomes (wheat and wild barley) were found. No meiotic pairing between chilense and turgidum chromosomes was detected. Differences in the meiotic behaviour per chromosome and amongst genomes are explained on the basis of cytomorphological and heterochromatin characteristics.     

Potential of Start Codon Targeted (SCoT) markers for DNA fingerprinting of newly synthesized tritordeums and their respective parents

Hexaploid tritordeum (H^chH^chAABB; 2n?=?42) results from the cross between Hordeum chilense (H^chH^ch; 2n?=?14) and cultivated durum wheat (Triticum turgidum ssp. durum (AABB; 2n?=?28). Morphologically, tritordeum resembles the wheat parent, showing promise for agriculture and wheat breeding. Start Codon Targeted (SCoT) polymorphism is a recently developed technique that generates gene-targeted markers. Thus, we considered it interesting to evaluate its potential for the DNA fingerprinting of newly synthesized hexaploid tritordeums and their respective parents. In this study, 60 SCoT primers were tested, and 18 and 19 of them revealed SCoT polymorphisms in the newly synthesized tritordeum lines HT27 and HT22, respectively, and their parents. An analysis of the presence/absence of bands among tritordeums and their parents revealed three types of polymorphic markers: (i) shared by tritordeums and one of their parents, (ii) exclusively amplified in tritordeums, and (iii) exclusively amplified in the parents. No polymorphism was detected among individuals of each parental species. Three SCoT markers were exclusively amplified in tritordeums of lines HT22 and HT27, being considered as polyploidization-induced rearrangements. About 70 % of the SCoT markers of H. chilense origin were not transmitted to the allopolyploids of both lines, and most of the SCoTs scored in the newly synthesized allopolyploids originated from wheat, reinforcing the potential use of tritordeum as an alternative crop.     

Construction and study of leaf rust-resistant common wheat lines with translocations of <Emphasis Type="Italic">Aegilops speltoides</Emphasis> Tausch. Genetic material

Genotyping was performed for the leaf rust-resistant line 73/00i (Triticum aestivum × Aegilops speltoides). Fluorescence in situ hybridization (FISH) with probes Spelt1 and pSc119.2 in combination with microsatellite analysis were used to determine the locations and sizes of the Ae. speltoides genetic fragments integrated into the line genome. Translocations were identified in the long arms of chromosomes 5B and 6B and in the short arm of chromosome 1B. The Spelt1 and pSc119.2 molecular cytological markers made it possible to rapidly establish lines with single translocation in the long arms of chromosomes 5B and 6B. The line carrying the T5BS · 5BL-5SL translocation was highly resistant to leaf rust, and the lines carrying the T6BS · 6BL-6SL translocation displayed moderate resistance. The translocations differed in chromosomal location from known leaf resistance genes transferred into common wheat from Ae. speltoides. Hence, it was assumed that new genes were introduced into the common wheat genome from Ae. speltoides. The locus that determined high resistance to leaf rust and was transferred into the common wheat genome from the long arm of Ae. speltoides chromosome 5S by the T5BS · 5BL-5SL translocation was preliminarily designated as LrAsp5.     

Development and characterization of wheat-<Emphasis Type="Italic">Ae. searsii</Emphasis> Robertsonian translocations and a recombinant chromosome conferring resistance to stem rust

The emergence of a new highly virulent race of stem rust (Puccinia graminis tritici), Ug99, rapid evolution of new Ug99 derivative races overcoming resistance of widely deployed genes, and spread towards important wheat growing areas now potentially threaten world food security. Exploiting novel genes effective against Ug99 from wild relatives of wheat is one of the most promising strategies for the protection of the wheat crop. A new source of resistance to Ug99 was identified in the short arm of the Aegilops searsii chromosome 3S^s by screening wheat- Ae. searsii introgression libraries available as individual chromosome and chromosome arm additions to the wheat genome. For transferring this resistance gene into common wheat, we produced three double-monosomic chromosome populations (3A/3S^s, 3B/3S^s and 3D/3S^s) and then applied integrated stem rust screening, molecular maker analysis, and cytogenetic analysis to identify resistant wheat-Ae. searsii Robertsonian translocation. Three Robertsonian translocations (T3AL·3S^sS, T3BL·3S^sS and T3DL·3S^sS) and one recombinant (T3DS-3S^sS·3S^sL) with stem rust resistance were identified and confirmed to be genetically compensating on the basis of genomic in situ hybridization, analysis of 3A, 3B, 3D and 3S^sS-specific SSR/STS-PCR markers, and C-banding. In addition, nine SSR/STS-PCR markers of 3S^sS-specific were developed for marker-assisted selection of the resistant gene. Efforts to reduce potential linkage drag associated with 3S^sS of Ae. searsii are currently under way.     

Inheritance and chromosomal locations of male fertility restoring gene transferred from Aegilops umbellulata Zhuk. to Triticum aestivum L.

Restriction fragment length polymorphism (RFLP) markers were used to map male fertility restoring gene that was transferred from chromosome 6U of Aegilops umbellulata Zhuk. to wheat. Segments of chromosome 6U bearing the gene that restore fertility to T. timopheevi Zhuk. male sterile cytoplasm were identified in all four translocation lines by two probes, BCD21 and BCD342. Lines 040-5,061-1 and 061-4 are T6BL.6BS6U translocations, while line 2114 is a T6AL.6AS-6U translocation. Line 2114 has a much larger 6U chromosomal segment and lower frequency of transmission of male gametes with the alien segment than the other three lines. The restoring gene carried by the 6U segment in 2114 showed high expressivity and complete penetrance. This restoring gene is designated Rf6. A homoeologous chromosome recombination mechanism is discussed for the alien gene transfer.Paper No. 823 of the Cornell plant breeding series