Androgenesis in Hexaploid Spring Wheat F2 Populations and Their Parents Using a Multiple-Step Regeneration System

The main goals of the wheat androgenesis experiment were to study the main phenomena of in vitro androgenesis in anther culture of F₂ populations (10) and their parents (6), to compare the usage of P-4 and C-17 media for the formation of embryo/callus and to demonstrate a new plant regeneration system. The P-4 induction medium was found to be significantly better than the C-17 in the number of responsive anthers (RA) and calli induced (CI) at the 1 % and 0.1 % level, respectively. Genotypic effect was evident in both RA and CI. The yields of F₂ populations in RA and CI were significantly higher than those of their parents regarding both media. The data confirmed the existence of heterosis for RA and CI in F₂ populations. The ratio of green/albino plant regeneration was more favourable in the C-17 derived embryo/calli than in the P-4 derived ones. The frequency of plantlet regeneration was enhanced in the group of unresponsive calli by the application of the multiple-step regeneration system. In this system the calli lacking well developed morphogenic structure were transferred to a new regeneration medium, containing a higher concentration of the same cytokinin, other cytokinin or basic medium, before the occurrence of irreversible changes in their physiology.     

Effect of parental genotypes and colchicine treatment on the androgenic response of wheat F1 hybrids

The effect of the parental genotypes and colchicine treatment on the androgenic response of wheat (Triticum aestivum L.) F1 hybrids was studied. For this, anthers from three F1 hybrids and their parents were cultured on W14 initiation medium and W14 supplemented with 0.03% colchicine. The number of responding anthers, microspore‐derived structures/100 anthers, green plants/embryos cultured, green plants/100 anthers and albino plants/100 anthers were recorded. It was observed that embryo formation and plant regeneration ability were genetically controlled and genotype dependent. In both treatments the variety Kavkaz had a significantly higher percentage of responding anthers, microspore‐derived structures and green plants/100 anthers than the other genotypes. On the other hand, the variety Myconos also demonstrated high microspore‐derived structure production and green plant regeneration when treated with colchicine. The good response observed in these two varieties indicates the importance of colchicine treatment only for certain genotypes. Green plant production capacity of the hybrids was intermediate to that of the parental varieties. As one parent with a high or even an intermediate response to anther culture could lead to the production of sufficient (for breeding purposes) green plants from the F₁ hybrids, it was concluded that screening the inbred lines for the response to anther culture with and without colchicine treatment could contribute to utilization of breeding material with a low response to anther culture via the proper hybrid combinations.     

利用Fhb1基因功能标记选择提高黄淮冬麦区小麦品种对赤霉病的抗性

赤霉病已上升为黄淮冬麦区的主要病害, 提高小麦品种对赤霉病的抗性成为该麦区主要的育种目标之一。宁麦9号、生选6号、建阳798、建阳84、苏麦3号和宁麦13均携带Fhb1基因, 对赤霉病表现中抗水平以上。本研究以这6个品种(系)为供体, 分别与高感赤霉病的周麦16矮败小麦近等基因系杂交和回交, 构建6个回交群体。利用Fhb1基因的KASP标记在回交后代中进行基因型分析, 分别选择携带和不携带Fhb1基因的可育株, 对后代株系进行单花滴注接种鉴定和田间病圃自然鉴定。回交后代携带Fhb1家系整体抗性达到中感, 比不携带Fhb1家系的平均病小穗数低4.2 (P < 0.01), 平均病情指数低4.0, 比轮回亲本周麦16的平均病小穗数和病情指数分别低8.1 (P < 0.01)和28.4 (P < 0.01)。不同供体品种(系)回交后代在赤霉病抗性上表现出明显差异, 以生选6号为供体的回交后代家系抗性表现最好。本研究表明, 利用Fhb1基因分子标记辅助选择技术能够有效地提高黄淮冬麦区小麦品种的赤霉病抗性水平。     

Selection for Fusarium head blight resistance in early generations reduces the deoxynivalenol (DON) content in grain of winter and spring wheat

Fusarium head blight (FHB) results in yield losses and contamination of kernels by mycotoxins, particularly deoxynivalenol (DON). For minimizing DON content in grain, indirect selection methods would increase gains from selection compared to the costly and time‐consuming DON analysis. The aim of this study was to examine whether an early selection for fewer FHB symptoms would lead to a reduced DON content in grain after inoculation with Fusarium culmorum. Starting with a double‐cross derived population of about 1,100 genotypes, 30 F_1:3 genotypes were selected for FHB rating in a two‐step selection in spring wheat with the non‐adapted resistance sources CM82036 and ‘Frontana’. In winter wheat, 30 F_1:2 genotypes were selected out of a double‐cross derived population of about 600 F₁ plants from crosses with German resistance sources (‘Dream’, G16‐92). Selected genotypes were grouped in three categories according to their FHB rating (low, moderate and high) and analysed afterwards for grain DON content. The three groups differed in their DON content illustrating that indirect selection should already be feasible in the earliest generations. Because of the wide genotypic ranges for DON contents within one grouping, a final DON analysis for selected materials is advisable to achieve full selection gain.     

Somaclonal Variation in Tissue Culture of Triticum aestivum×Agropyron desertorum F1 Hybrid

Callus induced from immature inflorescences of the partially self-fertile hybrids (2 n = 35; ABDPP) between Triticum aestivum (2n = 42; AABBDD) and Agropyron desertorum (2n = 28; PPPP) led to the regeneration of 88 plants on MS medium supplemented with 2 mg/l of 2,4-D. These regenerants were used to investigate somaclonal variation and to obtain more selfed derivatives. Immature inflorescences at the stage of developing floral primordia gave the best response in terms of callus induction and plant regeneration. The regenerants exhibited great variability for most morphological traits. Although the regenerants did not exhibit variation in chromosome number, they did show a higher degree of meiotic instability than the initial hybrid. In particular, the regenerants gave much higher selfed seed-set (5.49 %) than the donor hybrid (0.46 %), so that a total of 484 selfed seeds were obtained.     

Production of Doubled Haploids by Anther Culture and Wheat X Maize Method in a Wheat Breeding Programme

Utilization of the doubled haploid method of breeding usually shortens the time to cultivar release, and methods of haploid production need evaluation in a breeding programme. Thirty-eight different three-way crosses were tested for anther culture response. On average 5.8 percent of the anthers cultured produced calli. Three crosses were found recalcitrant for callus induction. Overall, the anther culture method produced 0.6 plantlet per 100 anthers cultured. Five crosses with an average of 5.8 and 2.8 percent of anthers producing calli and plantlets, respectively, were compared using anther culture and wheat × maize crosses. Non-responsive genotypes for callus induction and plantlet formation in the anther culture method proved to be good parental material in wheat × maize crosses. The average percentages of embryo formation and plantlet production in wheat × maize crosses were 10.3 and 4.7, respectively. Anther-derived plants were cytologically unstable, whereas all the plants regenerated from wheat × maize crosses were haploids (n = 21 chromosomes). The chromosome numbers of the polyhaploids were doubled with a colchicine treatment. Improvement of the two haploid production methods to facilitate their efficient use in a breeding programme is discussed.     

Anther culture response of F1 durum × bread wheat hybrids after colchicine application

The effect of colchicine on the androgenic response of durum × bread wheat F₁ hybrids was studied. For this, three Greek durum wheat cultivars, exhibiting good yield and pasta quality, were crossed to two bread wheat cultivars with good response to anther culture. Spikes of the resulting hybrids, containing microspores in the mid to late uninucleate stage, were selected and cultured on two different media (W₁₄ and solid potato‐2) with and without colchicine. A negative effect on anther culture response was observed when colchicine was added to the W₁₄ induction medium. A similar effect was observed in two of the hybrids when they were cultured on to potato‐2 induction medium. In both induction media, green plant production was influenced negatively by colchicine treatment. The same was observed in albino plant production. The results of the present study support the view that anther culture response is strongly genotype dependent. Finally, potato‐2 induction medium was the most suitable one for the material studied.     

Potential use of doubled haploid lines for the screening of resistance to yellow rust (Puccinia striiformis) in hexaploid wheat

Doubled haploid lines derived from anther culture of two Iranian spring wheat genotypes‘Ghods’susceptible and‘9106’resistant to yellow rust in Iranian field conditions, and their F₁ hybrids were used in this study. Seedlings of 36 doubled haploid lines, selected out of 96 according to their agronomic traits and the two parental genotypes were inoculated with eight races of yellow rust. The parental genotypes (‘Ghods’and‘9106’) were segregating for some of the races but their doubled haploid lines were either resistant or susceptible to them.‘Ghods’was susceptible to three of the races studied but three doubled haploid lines derived from it were resistant to them. Five selected doubled haploids from the‘9106’genotype and six from F₁ hybrid plants were resistant to all eight races tested. After further investigations in Iranian field conditions it was found that some of these lines can be used as donor genotypes for resistance to yellow rust in wheat breeding programmes. Use of these genotypes should be possible if the French yellow rust races used for selection also represent the dominant races in Iran. It can be concluded that anther culture provides an efficient method for fixing genes of resistance to yellow rust and desirable doubled haploids from F₁ plants can be derived.     

Efficient application of in vitro anther culture for different European winter wheat (Triticum aestivum L.) breeding programmes

The efficiency of our anther culture protocol was tested with high‐ and low‐responding genotypes, ‘Svilena’ and ‘Berengar’, and 93 F₁ winter wheat crosses in 2010 and 2011. Based on data for these genotypes, the effect of genotype influenced the number of embryo‐like structures, regenerated plantlets and green plantlets, while the number of albino plantlets was affected by genotype, year and environmental factors. Although genotype also influenced the production of green plantlets from breeding crosses, with green plantlets per 100 anthers ranging from 0.04 to 28.67, the average regeneration rate over all crosses was 5.3 green plantlets/100 anthers, which resulted in a total of 11 416 well‐rooted green plantlets. The survival rate of green plantlets following acclimatization was 97.21% in 2010 and 96.34% in 2011. In this study, the phenomenon of albinism and genotype dependency did not hinder the production of more than five thousand green plantlets each year. In our experiments, anther culture proved to be an efficient method in winter wheat breeding programmes with lower costs than alternative technologies.     

Physiological Genetics of Salt Tolerance in Wheat (Triticum aestivum L.): Performance of Wheat Varieties, Inbred Lines and Reciprocal F1 Hybrids under Saline Conditions

Four bread wheat cultivars were studied at two salinity levels. Tobari 66 had the lowest uptake of Na⁺ and Cl⁻, and the highest K⁺/Na⁺ ratio; Pato had the highest uptake of these ions and Lyallpur 73 was intermediate. Intervarietal differences were greater at higher salinity, suggesting that they were not caused by variation at the Kna1 locus. There were significant differences between inbred lines for Na⁺, particularly in Blue Silver, suggesting the possibility of selecting genotypes with enhanced tolerance from within existing cultivars. Pato, Tobari 66 and their reciprocal F₁ hybrids were further evaluated at four salinity levels. The hybrids exhibited similar relative grain yield to Tobari, with better Na⁺ and Cl⁻ exclusion and higher K⁺/Na⁺ ratios than Pato. Overall, Tobari had the highest absolute yield under salinity, and the hybrids were closer to Tobari than to Pato. Tiller and grain numbers, 100-grain weight and yield were more affected by salinity than were height, spike length and spikelet number. We conclude that intervarietal variation for salt tolerance in wheat is controlled by genes which could be transferred to sensitive genotypes to improve their tolerance, and that the K⁺/Na⁺ ratio of the youngest leaf could be used to screen for salt tolerance.     

Anther-culture response in different genotypes and F1 hybrids of pepper (Capsicum annuum L.)

An in vitro anther-culture method has been improved by using young mother plants and by using frequent subcultures, thus increasing the androgenic yield in different Capsicum annuum L. genotypes. An assortment of peppers was used, composed of 15 genotypes (four breeding lines, seven cultivars and four F₁ hybrids). A new system for qualifying the androgenic response was established. For use in practical breeding, a minimum of 5 % of plant regeneration was proposed as the criterion for a fair response. Accordingly, one excellent, one good and eight fair responses were identified among the genotypes investigated. As compared to the standard cultivar. 2 genotypes gave a significantly better response, i.e. ‘Fehérözön’ (75.8%) and ‘Szechuan 90716’ (21.0%). In comparative investigations, F₁ hybrids, produced from crosses between poor/non-responsive and responsive genotypes, showed a fair level of response, even the case of a poor response in donor parent. The ploidy level of the resulting plants was determined by flow-cytometric analysis.     

Evaluation of F2 and F3 plants introgressed with QTLs for clubroot resistance in cabbage developed by using SCAR markers

Clubroot disease caused by Plasmodiophora brassicae is one of the major diseases of Brassica crops, often devastating to the cultivation of cruciferous crops in temperate regions. In a previous study (Moriguchi et al. 1999) identified three major quantitative trait loci (QTLs) for clubroot resistance, each in a separate linkage group, in a population derived from a cross between a clubroot‐susceptible inbred cabbage line, Y2A and a resistant inbred kale line, K269. In this study, the original random amplified polymorphic DNA (RAPD) and restriction fragment length polymorphism (RFLP) markers were converted into sequence‐characterized amplified region (SCAR) markers to facilitate large‐scale marker‐assisted screening of clubroot resistance in cabbage breeding. Of 15 RAPD markers closely linked to the three QTLs, nine SCARs were developed as dominant markers after cloning and sequencing. In addition, two RAPD markers were converted into co‐dominant cleaved amplified polymorphic sequence (CAPS) markers, and one RFLP marker out of three tested was converted to a dominant SCAR marker. The effect of selection for resistance by the improved markers was evaluated in progeny plants in the F2 and F3. A total of 138 F2 plants were genotyped with nine SCARs and 121 well‐distributed makers consisting of 98 RAPD, 19 RFLP, two isozymes, and two morphological markers in order to estimate the level of resistance and the proportion of undesirable alleles from the kale in non‐target areas in each of the F2 populations. An F2 plant, YK118, had kale alleles at QTL1, QTL3 and QTL9. Three F2 plants, namely, YK107, YK25 and YK51 had kale alleles at only QTL1, QTL3 and QTL9, respectively. These F2 plants were selected for their low proportion of alleles derived from kale in non‐target regions. YK118, like the resistant kale parent, expressed very high resistance to three field isolates of Plasmodiophora brassicae, whereas the mean disease index in the F2 and F3 plants carrying only single QTLs was intermediate. The QTLs showed no differential response to the isolates. These plants with improved resistance will be useful as parental inbred lines for F1 hybrids.     

Medium-Genotype-Interaction on Androgenetic Haploid Production in Wheat

The present study investigates the effects of media on the rate of green plantlet formation from anther cultures of five different winter and spring wheat cultivars. An increased number of embryos and calli were produced on Ficoll-containing liquid potato-2 medium, whereas the addition of 0.2 M/l maltose increased the rate of regeneration. The genotypes had strong effect on the formation of plantlets, but the developed procedure allows also the regeneration of recalcitrant genotypes. On the best medium sequence, a total of 2430 anthers over all genotypes were plated which developed 1325 macroscopic structures (55 %) of which 159 green plants have developed, i.e. 12 % from the structures and 7 % from the plated anthers, respectively.     

Relationship of Root and Shoot Characters in Parent, F1 and F2 Populations of Rice

The direct and indirect contributions of root characters — root length, roots/plant, fresh and dry root weight on grain yield/plant were worked out from a 7 × 7 diallel set of rice hybrids. The materials were grown in pots with four replications. Path analysis was done at genotypic level of correlation.
The grain yield/plant showed positive correlation with all the root characters in parent, F₁ and F₂ except with roots/plant in F₁ population. Fresh root weight demonstrated positive direct effect on grain yield/plant in all the three generations. Roots/plant had highly positive direct effect in F₂. Direct effects were negative in respect of root length and dry root weight in F₂ generation.     

Efficiency of Early Selection for Yield Performance in Wheat

Selection for yield in early generations is difficult, mainly due to a shortage of seed. In the breeding company Fr. Strube the breeder uses three-way crosses and first selects for yield between F₂′-derived lines in the F₄′. The generations are called F₁, F₂′, etc., because the first generation is already segregating. The efficiency of this selection was checked in the F₅′ and F₆′. For this purpose F₃′-derived progenies, corresponding to poor and good yielding bulks in F₄′, were selected. In later generations the average yield of the group of strains with good yield in the F₄′ was 4 % (F₅′) and 3 % (F₆′) higher, respectively, than the mean of the strains with low yield in the F₄′. Strains with very low yielding potential can easily be discarded in the F₄′. The risk of discarding lines with a very high yield is minimal. Selection for yield in the F₄′ was as effective as in the F₅′. The importance of testing F₂′-derived bulks in the F₄′ is not only based on the assessment of yield but also on the possibility of reliable intensive selection through recording visual characters (height, resistance against diseases) in a second environment in parallel to the single plant progenies.     

Variation and covariation for quantitative resistance to head blight (Fusarium culmorum) in two testcross series of S2 lines in winter rye

The amount of genetic variation among inbred lines and testcrosses, and covariation between both genetic materials, are of crucial importance for selection efficiency in hybrid breeding. To estimate these quantitative genetic parameters for resistance of winter rye (Secale cereale) to head blight caused by Fusarium culmorum, 88 three-way cross hybrids, produced by crossing each of 44 S₂ Carsten inbred lines with two unrelated Petkus single-cross testers, were evaluated along with the parental lines over 2 years. Resistance traits were head-blight rating and grain weight per spike relative to the non-inoculated control. Significant genotypic variation occurred among lines and in both testcross series. S₂ lines displayed considerably more variation than testcross series. Genotype × environment interaction was more marked among the inbred lines, while estimates of heritability were similar for both genetic materials. Testcrosses showed heterosis for head-blight resistance. No relationship existed between S₂ lines and the two testcross series for any resistance trait. This might be caused by an association between inbreeding and Fusarium-head-blight susceptibility and different inbreeding depression among the S₂ population. The phenotypic correlations between the testcross series were moderate for both traits (r = 0.58, P < 0.01). In conclusion, Fusarium-head-blight resistance has to be selected at the respective heterozygosity levels.     

Large-scale production of wheat and triticale double haploids through the use of a single-anther culture method

A total of 257 parental wheat and 38 triticale lines were used for anther culture. On average, 2.1 green wheat haploids were obtained per spike. This response occurred irrespective of the origin of the material (Germany, France, Sweden or UK) and 5 years of testing. Triticale responded with 5.3 green haploids per spike. Using the criterion that one parental line should give at least one green haploid per spike in the screening experiment, green haploids were produced from 88 out of 91 F₁ wheat breeding combinations and from each of 21 F₁ and F₂ triticale breeding combinations. An average of 4.7 green plants were obtained per spike from the wheat production programme, while the triticale programme gave an average of 6.2 green plants per spike. A single medium supplemented with different hormones for anthers and embryos was used for culture of both crops.     

Types and components of resistance to Fusarium head blight of wheat

Resistance of wheat to Fusarium head blight caused by Fusarium graminearum and F. culmorum was identified in natural epidemics in 1985 and 1987 as well after artificial inoculations (1983–1988 and 1984–1987). Out of 25 genotypes tested, five were identified with no significant difference in head blight scores, but differing significantly in yield after artificial inoculation, i.e. tolerance differences were detected at different resistance levels. Some genotypes that were similar in yield or head blight scores differed in seed infection severity. Genotypes with awns were more susceptible to head blight when tested under natural epidemic condition in the field; but this trait did not influence head blight severity in artificial inoculations. Dwarf genotypes were more severely infected by head blight than tall genotypes under natural conditions, but genotypes of different plant height classes were similarly susceptible after artificial inoculations. In the early generations of a breeding programme resistance measured by visual evaluation of artificial inoculation is the most important way to screen. If selection of dwarf and awned genotypes cannot be avoided, the higher susceptibility caused by awns and dwarfness under natural epidemic conditions can be decreased by a higher level of physiological resistance, as variability in physiological resistance is available. In later generations, traits like percentage of seed infection or tolerance can be identified by additionally measuring yield reduction. Stability of disease reaction appears to be connected with resistance level, the most resistant genotypes are the most stable, and the most susceptible ones tend to have more unstable reactions in different epidemic conditions.     

Phenotypic selection for high resistance to Fusarium head blight after introgression of quantitative trait loci (QTL) from exotic spring wheat and verification by simple sequence repeat markers a posteriori

Fusarium head blight (FHB) has become an important disease of wheat. We introgressed three resistance quantitative trait loci (QTL) alleles on chromosomes 3B, 5A (from CM82036) and 3A (from ‘Frontana’) into European elite spring wheat and performed phenotypic selection among double‐cross (DC) derived progeny in generations DCF₂ and DCF₃. After recombination and selfing, we analysed 135 phenotypically selected progeny by simple sequence repeat (SSR) markers linked to the QTL. In a second experiment, we forwarded the best 20 progeny for a further two generations by pedigree selection. Progeny were inoculated at two to four locations with Fusarium culmorum and the percentage of infected spikelets per plot was estimated. Both experiments show that phenotypic selection was highly effective. One‐hundred out of 135 phenotypically selected DCF_1:3 progeny had the combination of donor‐QTL alleles (3B + 5A + 3A, 3B + 5A) with the highest effects on FHB resistance. In the subsequent generations, sufficient genotypic variance was detected. The best F_5:7 bulks had similar resistance to the donor CM82036. The FHB rating was reduced in total by 45% points compared to the parental mean. QTL with high effects can be detected solely by phenotypic selection after targeted introgression.     

QTL mapping and marker-assisted selection for Fusarium head blight resistance in wheat: a review

During the past decade, numerous studies have been published on molecular mapping of Fusarium head blight (FHB) resistance in wheat. We summarize the relevant findings from 52 quantitative trait loci (QTL) mapping studies, nine research articles on marker-assisted selection and seven on marker-assisted germplasm evaluation. QTL for FHB resistance were found on all wheat chromosomes except chromosome 7D. Some QTL were found in several independent mapping studies indicating that such QTL are stable and therefore useful in breeding programmes. We summarize and update current knowledge on the genetics of FHB resistance in wheat resulting from QTL mapping investigations and review and suggest FHB breeding strategies based on the available information and DNA markers.