Marker‐assisted backcross breeding to combine multiple rust resistance in wheat

A widely grown but rust susceptible Indian wheat variety HD2932 was improved for multiple rust resistance by marker‐assisted transfer of genes Lr19, Sr26 and Yr10. Foreground and background selection processes were practised to transfer targeted genes with the recovery of the genome of HD2932. The near‐isogenic lines (NILs) of HD2932 carrying Lr19, Sr26 and Yr10 were individually produced from two backcrosses with recurrent parent HD2932. Marker‐assisted background selection of NILs with 94.38–98.46% of the HD2932 genome facilitated rapid recovery of NILs carrying Lr19, Sr26 and Yr10. In the BC₂F₂ generation, NILs were intercrossed and two gene combinations of Lr19+Yr10, Sr26 + Yr10 and Lr19+Sr26 were produced. A total of 16 progeny of two gene combinations of homozygous NILs of HD2932 have been produced, which are under seed increase for facilitating the replacement of the susceptible HD2932 with three of the sixteen improved backcross lines with resistance to multiple rusts.     

Resistance to multiple leaf spot diseases in wheat

Tan spot (TS), Stagonospora nodorum blotch (SNB), and Septoria tritici blotch (STB) are three major leaf spot diseases of wheat worldwide. Host plant resistance (HPR) is one of the main components in the management of these diseases in wheat. The objective of this study was to identify new sources of resistance to TS (races 1 and 5), SNB, and STB. A total of 164 wheat genotypes developed by the International Maize and Wheat Improvement Center (CIMMYT), Mexico were individually evaluated for TS, SNB and STB in spring and fall of 2006 in the greenhouse. Two experiments were conducted in a randomized complete block design with three replicates. Each replicate consisted of 164 wheat genotypes planted in cones with three seedlings/genotype in each cone and disease reaction was assessed for each race or pathogen at the two- to three-leaf stage. Based on the disease reactions, three wheat genotypes were resistant to both TS and SNB, while 13 genotypes were resistant to TS and STB. Similarly, 13 genotypes were resistant to both SNB and STB. In addition, four wheat genotypes were highly resistant to TS, SNB, and STB. These results suggest that the resistant genotypes identified in this study possess high levels of resistance to multiple leaf spot diseases and could be valuable sources for wheat improvement programs.     

Spontaneous hybridization in wheat populations of Transcaucasia

Summary Hybrids naturally arising between Triticum species and between Triticum species and species of Secale and Aegilops are found in wheat fields in Transcaucasia. This continuous exchange of genetic material will yield new genotypes which may be useful in breeding.This active source of the origination of new botanical (sub)varieties results in a great diversity of species and varieties which led Vavilov to conclude that Transcaucasia is centre of origin of several Triticum species and varieties.     

Fluorescent in situ hybridization as an aid to introducing alien genetic variation into wheat

Summary Fluorescent in situ hybridization (FISH) has been used to assess the occurrence and frequency of wheat-alien chromosome pairing in a wheat/Thinopyrum bessarabicum hybrid and in wheat/rye hybrids with different levels of chromosome pairing by examining pollen mother cells at metaphase I of meiosis. The use of FISH to identify the presence and size of alien chromatin in a wheat background is also demonstrated.The value of FISH as an aid to the introgression of alien genetic variation into wheat is discussed.Abbreviations FISH fluorescent in situ hybridization - GISH genomic in situ hybridization - PRINS primer-induced in situ hybridization     

Pollen sterility induced by ethrel and its utilization in hybridization of wheat

Summary In 1974 and 1975 gametocidal effects of Ethrel were investigated under field conditions in the spring wheat cv. Zlatka. In experimental plants grain setting per spikelet and ear as well as ability to be cross-pollinated were ascertained.As compared with the control, treated variants achieved more than 90% sterility. Female fertility was not affected and cross-pollination was possible. Genetical check showed that the best variants, when cross-pollinated, produced 50–55% hybrids; grain setting, however, was brought down by more than 37%. As a result of Ethrel treatment haulm length was reduced and the vegetation period was somewhat prolonged.     

Searching for DNA introgressed from wheat and for wheat-like grain proteins in a rice x wheat hybridization derivative

The DNA of a putative rice x wheat hybridization derivative (X Oryticum oryzoides) from China, the DNA of its parental rice cultivar and the DNA of a wheat line were digested with ten different restriction endonucleases, resolved by agarose electrophoresis, Southern blotted and hybridized using genomic wheat DNA as a probe. Phenol extracted, ethanol and cetyltrimethylammonium bromide precipitated DNA of the putative hybrid showed a restriction fragment length polymorphism (RFLP) different from that of the parental rice. When the DNA was further purified by Qiagen chromatography, the RFLP differences were not detected. Hence the apparent RFLP differences were probably due partial digestion of the less pure DNA preparations by the restriction endonucleases. No real introgressed fragments from wheat genome could be shown. The HpaII/MspI sites were more frequently digested with MspI than with HpaII in rice and hybridization derivative DNA, but the sites were evidently more frequently methylated in wheat DNA. Thus, in terms of methylation of the DNA, the hybridization derivative was much more like the rice parent than the wheat parent. The hybridization derivative showed a single endospermal protein (mass 19 kg mol^-1) not detected in the parental rice cultivar. This minor protein was soluble in buffered 50% isopropanol and precipitable with methanol. The results indicate that there are no or only short introgressed sequences from wheat in the rice/wheat derivative, a result which might be considered in breeding efforts with the hybrid derivative.     

Fluorescent in situ hybridization — a useful aid to the introduction of alien genetic variation into wheat

Summary Fluorescent in situ hybridization (FISH) of DNA to plant chromosomes has proved to be a powerful cytogenetic tool. The value of fluorescent in situ hybridization of total genomic DNA (GISH) of related species is demonstrated in the determination of wheat/alien chromosome pairing in hybrids. Its use for assessing the relative merits of the various genes that affect chromosome pairing is also shown.The ability of GISH to identify the presence in wheat of whole alien chromosomes or alien chromosome segments is illustrated. The potential of FISH for detecting repeated DNA sequences, low copy sequences and single copy genes is discussed.Abbreviations FISH fluorescent in situ hybridization - GISH genomic in situ hybridization - PRINS primer-induced in situ hybridization     

Wide hybridization between wheat (Triticum L.) and lymegrass (Leymus Hochst.)

A total of 240 F₁ hybrids was made beween wheat (Triticum aestivum L. em. Thell. (2n = 6x = 42) and T. carthlicum Nevski (2n = 4x = 28)) and perennial lymegrass (North European Leymus arenarius (L.) Hochst. (2n = 8x = 56) and North American L. mollis (Trin.) Pilger (2n = 4x = 28)). The wide crosses yielded embryos in 20% of caryopses and 96% of the embryos developed into normal hybrid plants. The hybrids were vegetatively vigorous, with evidence of the Leymus rhizomatous habit. Those deriving from L. arenarius survived overwintering in Iceland, but the hybrids L. mollis did not, whereas in a milder environment, both showed perenniality. Cytogenetic analysis of root tip cells before the plants were treated with colchicine showed that 21 out of 28 hybrids investigated had chromosome mosaics, with a population of both amphihaploid and amphidiploid cells. This spontaneous doubling of somatic chromosomes occurred in all cross combinations, with the highest average frequency of diploid cells (28%) in T. carthlicum × L. arenarius crosses. A few selfed seeds have been obtained from a T. aestivum × L. arenarius hybrid. All the hybrids were treated twice with colchicine, but the treatment appeared to have little or no effect on the frequency of chromosome doubling in the hybrids deriving from T. aestivum. The frequency of diploid cells, however, increased significantly (e.g. to 80%) in the hybrids deriving from the T. carthlicum parent. Genomic in situ hybridization confirmed the hybridity of the plants and showed that the hybrids were amphiploids containing genomes of both wheat and lymegrass. In situ hybridization using ribosomal DNA probe differentiated chromosomes of L. mollis, L. arenarius from those of wheat. The hybrids are being backcrossed with lymegrass pollen, aiming to domesticate the wild, perennial species. This revised version was published online in July 2006 with corrections to the Cover Date.     

Fluorescence in situ hybridization polymorphism using two repetitive DNA clones in different cultivars of wheat

Twenty‐two wheat cultivars and a wheat line were analysed with two‐colour fluorescence in situ hybridization (FISH) using the pSc119.2 and pAs1 repetitive DNA clones to detect if polymorphism could be observed in the hybridization patterns of different wheat cultivars. The FISH hybridization pattern of ‘Chinese Spring’ was compared with wheat cultivars of different origins. Differences were observed in the hybridization patterns of chromosomes 4A, 5A, 1B, 2B, 3B, 5B, 6B, 7B, 1D, 2D, 3D and 4D. Although a low level of polymorphism exists in the FISH pattern of different wheat cultivars, it is possible to identify 17 pairs of chromosomes according to their hybridization patterns with these two probes. This study will help to predict the expected variation in the FISH pattern when analysing wheat genetic stocks of different origin. It is presumed that variation in hybridization patterns are caused by chromosome structural rearrangements and by differences in the amount and location of repetitive sequences in the cultivars analysed.     

Simulations and practical problems of applying multiple marker assisted selection and doubled haploids to wheat breeding programs

Marker assisted selection (MAS) and wheat doubled haploids (DH) are relatively new technologies, recently applied to wheat breeding programs. Simulations demonstrate that DHs increase the efficiency of MAS, and offer faster strategies for combining large numbers of genes with a minimum number of marker tests. When small numbers of marked loci (1-3) are selected simultaneously, selection of DH progeny is 5-6 times more efficient than selecting F₄ derived families. Combining 4-8 marked loci, screening of F₂ plants and using only those plants homozygous or segregating for all of the marked loci as parents for DH production (10-31% of F₂ plants) is 3-10 times as efficient as using F₁ plants. A number of protocols have been proposed involving sib-matings and selection to fix some genes, with further selection in the second generation to improve the proportion of useful DH lines. In one scheme (recombinant F₂ selection) all F₂ plants, either homozygous or heterozygous for the marked alleles, are intercrossed at random and the recurrent F₁ plants still having these alleles are used for DH production. An alternative strategy (recurrent DH selection) is to select from an initial DH population and intercross those lines having most favourable marked loci with a second cycle of DHs to fix all favourable marked loci. Combining more than 12 marked gene loci does not seem feasible, due to the very large numbers of F₂s (>2000) required. This has implications when using MAS for quantitative trait loci, where many minor gene loci would have to be combined. Direct selection for some multi-genic quantitative traits amongst the DH lines may be more efficient than using MAS where recurrent selection is used. At the Cereal Research Centre, the practical problems of using these protocols as part of the spring wheat breeding program are being evaluated. This revised version was published online in August 2006 with corrections to the Cover Date.     

Sequential combinations of C-banding and in situ hybridization and their use in the detection of interspecific introgressions into wheat

Summary The sequential combination of C-banding and in situ hybridization techniques applied in this or in a reverse order, are used to recognize targeted chromosomal regions in cereals. Both methods are described whereby standard chromosome squash preparations are followed by: i) C-banding technique using Leishman stain and a slightly modified in situ hybridization technique using biotin-labeled DNA probes, or ii) fluorescence in situ hybridization technique and C-banding. Both approaches have been successfully used onto mitotic chromosomes of rye and wheat resulting suitable for both their identification and detection of targeted sites.     

The use of microsatellite markers to determine the relative proportions of grain produced by cultivars and the frequency of hybridization in bread wheat mixtures

Two polymorphic microsatellite markers were selected to identify 24 bread wheat cultivars commonly grown in France and to estimate the proportions of cultivar and hybrid grains in the harvests of four 4‐cultivar mixtures (CM 1‐4) planted in equal proportions in farmers’ fields. This technology was used not only to determine whether a mixture comprises the declared cultivars, but also whether there is any contamination with other cultivars, to identify the contaminants and to estimate their proportions. At harvest, the cultivar proportions ranged from 15 to 37%. Only in the mixture CM2 (‘Malacca’, ‘Somme’, ‘Renan’ and ‘Soissons’), did the cultivars contribute equally to the harvest. The other cultivar mixtures CM1 (‘Malacca’, ‘Somme’, ‘Renan’ and ‘Camp Remy’), CM3 (‘Malacca’, ‘Texel’, ‘Apache’ and ‘Aligre’) and CM4 (‘Malacca’, ‘Somme’, ‘Apache’ and ‘Virtuose’) showed significantly unequal cultivar proportions with ‘Somme’ dominating ‘Renan’ and ‘Camp Remy’ in CM1, and ‘Apache’ dominating ‘Malacca’ and ‘Aligre’ in CM3 and ‘Malacca’ in CM4. Similar cultivar proportions were measured with gliadin and glutenin markers in the mixtures CM3 and CM4, confirming the results with microsatellites. No contamination was found. Hybrids accounted for between 1.3 and 6.3% of the grains produced in the four cultivar mixtures.     

Barriers to interspecific hybridization in Ulmus americana

Summary Crossability barriers between the tetraploid Ulmus americana and five other diploid elm species were investigated using both fluorescence and scanning electron microscopy. Both pollen germination and pollen tube development were inhibited in all interspecific crosses. Pollen tube behavior on the stigma of U. americana was unchanged when pollen of a colchicine-induced U. pumila tetraploid was substituted for diploid U. pumila pollen.Self-incompatibility was observed in one of two American elms studied and manifested itself via reduced pollen germination and inhibited pollen tube growth on the stigma surface.The results indicate that crossing barrier(s) between U. americana and other elm species occur at the stigma surface and operate independently of ploidy level. This conclusion is discussed in relation to current breeding efforts to transfer Dutch elm disease resistance into the American elm via interspecific hybridization with Eurasian elms.     

Identification for H. villosa chromatin in wheat lines using genomic in situ hybridization, C-banding and gliadin electrophoresis techniques

Detection of H. villosa chromosomes in telosomic addition and translocation lines of common wheat was undertaken using genomic in situ hybridization (GISH), C-banding techniques and polyacrylamide gels electrophoresis. The result of GISH on mitotic metaphase cells of the addition line `95039' indicated that the added telochromosomes originated from H. villosa, and it was probably 6VS or 7Vs of H. villosa according to the C-banding pattern. Furthermore, the analysis of gliadin profiles demonstrated that the telochromosome was 6VS. A pair of 1RS/1BL translocation chromosome was also found in `95039'. In addition, mitotic GISH analysis showed that the 6VS/6AL translocation chromosome remained unchanged after being transferred into new wheat background. This revised version was published online in August 2006 with corrections to the Cover Date.     

Genetic and environmental variability in wheat characteristics reported to be involved in morphological resistance to wheat stem rust

Summary Four morphological characteristics of wheat peduncles were evaluated for genetic and environmental variability. The thickness of the epidermis and the amount and distribution of chlorenchyma tissue in the peduncle were measured. The genetic variability was estimated to be large, whereas the environmental variability was small. It was also concluded that cultivars could be screened for morphological characteristics of the peduncle that contributed to non-specific resistance to stem rust with a minimum sample size of about 10 plants per replication from a single location.Paper No 9328, Scientific Journal Series, Minnesota Agricultural Experiment Station. University of Minnesota. St. Paul. MN 55108.     

Partial resistance to wheat leaf rust in 18 spring wheat cultivars

Summary Eighteen spring wheat cultivars were tested in microfields and race nurseries for their partial resistance PR to wheat leaf rust under low and high disease pressure respectively. Large differences existed between the 18 cultivars, Skalavatis 56 being the most susceptible and Ponta Grossa 1 being the most resistant cultivar. Of the three epidemic parameters, disease severity (DS) at the time that the susceptible check was severely diseased and area under the transformed disease severity curve (AUTC) and the logistic growth rate (r), AUTC and DS were highly correlated. Both seemed to be reliable estimators of PR but DS should be preferred for economical reasons. The logistic growth rate seemed to be unsuitable as an estimator of partial resistance.High and low disease pressure gave similar cultivar ranking. PR can be screened and selected equally well in race nurseries with low space, low time and low cost input as in microfields with high space, time and cost input.Cultivar differences in development rate had a large impact on the cultivar differences for amount of disease and can therefore greatly bias the estimation of cultivar resistance. The resistance of early cultivars tended to be underestimated whereas the resistance of late cultivars tended to be overestimated. The effect of differences in developmental rate was most pronounced in the flag leaf. It is advisable to avoid the assessment of disease levels on the flag leaf only and to incorporate in the tests several susceptible and resistant checks that cover the range of development rates in the material to be selected, because otherwise selection for resistance will tend to select also for lateness.Regression of the epidemiological parameters on three components of partial resistance revealed that latency period (LP) is an important factor in determining the resistance observed in the field explaining on average 67% of the observed variation. Adding infection frequency (IF) and urediosorus size (US) to the linear model increased the proportion of the observed variation in the field explained by the components to 80%. This result supports the idea that the components of PR inherit independently, at least, in part.     

Adaptation of wheat rusts to the wheat cultivars in former Czechoslovakia

Summary In former Czechoslovakia virulence of rusts attacking wheat was studied since the sixties. Since the same time genes for resistance in the registered cultivars were identified. The role of Berberis and Thalictrum as alternate hosts for stem rust and leaf rust, respectively, was investigated as well. Determined changes of virulence in rust populations could only partially be ascribed to changes of resistance genes in the grown cultivars. Unnecessary genes for virulence had no negative effect on the fitness of the pathogen. All tested samples of aeciospores from barberries attacked rye, not wheat. None of Thalictrum species occurring in the Czech and Slovak Republics was found to host wheat leaf rust. However, the sexual stage of wheat stem rust and wheat leaf rust could be induced on Berberis vulgaris and Thalictrum speciosissimum, respectively. General epidemiological conclusions are drawn from the results and experience of the last 35 years.