Wheat Culture Research Articles

Regeneration of plantlets from unpollinated ovary cultures of Ethiopian wheat (Triticum turgidum and Triticum aestivum)

An in vitro culture protocol was established for direct regeneration of plantlets from unpollinated ovary cultures of four Ethiopian wheat varieties. Unpollinated ovaries were excised from durum wheat (Yerer and Ude varieties) and bread wheat (Simba and Galama varieties). Analysis of variance (ANOVA) has shown that genotypes, types of media, concentrations of 2,4-dichlorophenoxy acetic acid (2,4-D) and kinetin (KIN) and durations of cold pretreatment at 4°C significantly (P≤0.05) affected direct formation of embryonic tissues independently. Stage II of wheat spikes, MS medium containing 1 mg/l of each of 2,4-D and KIN and 15 days of cold pretreatment were found to be the best conditions for direct formation of embryonic tissues. The highest frequency of shoots were regenerated from the cultured embryonic tissues of Yerer (41.6% ) and Simba (41.3%) on medium containing 0.1 mg/l 2,4-D. From a total of 14,524 cultured unpollinated ovaries, 1,100 embryonic tissues (7.6 %) and 75 regenerants were obtained. The average percentage of embryonic tissues and regenerants were 9.0 and 1.1% from 3,444; 9.8 and 0.55% from 4,732; 5.6 and 0.17% from 2,988; 4.7 and 0.12% from 3,360 cultured unpollinated ovaries for varieties Yerer, Simba, Ude and Galama, respectively. Keywords: Embryonic tissues, Unpollinated ovaries , regenerants, wheat varieties African Journal of Biotechnology Vol. 12(39), pp. 5754-5760

Effect of Cold-Mediated Pretreatment on Microspore Culture in Winter and Spring Wheat

Microspore culture of wheat generates completely homozygous (doubled haploid) plants in a single generation thereby reducing the time required for wheat variety development. Success of microspore culture in spring wheat is relatively higher than that in winter wheat. Cold mediated pretreatment was reported to improve response of microspore culture in wheat. The objective of the study was to determine and compare the influence of cold pretreatment on microspore culture in spring and winter wheat. Three spring (“Chris”, “Express”, and “Macon”) and three winter (“Anton”, “Antelope”, and “Camelot”) wheat cultivars were used. In cold pretreatment, excised anthers were incubated in solution B at 25°C-28°C for 4-5 days followed by cold treatment at 4°C for 5 days and were compared with the no-cold pretreatment at 25°C-28°C for 4-5 days. Isolated microspores were cultured in induction medium (MMS4) at 27°C-28°C for 25-30 days in the dark. Embryos (1-2 mm size) were transferred to regeneration medium (MMS5). Numbers of multicellular structures, transferable embryos and green plants were counted and data were used for analysis of variance using a generalized linear model. It was observed that cold pretreatment increased multicellular structures, transferable embryos and green plants in both spring and winter wheat. However, the degree of improvement was higher in spring wheat compared to winter wheat. The cultivars within spring and winter wheat responded differently. Development of embryos from pro-embryos was 4-5 folds lower in winter wheat than that in spring wheat, indicating requirement of a possibly different hormonal composition in induction medium for improving embryo induction in winter wheat. This report may provide future direction of research to improve microspore culture response in winter wheat.

Major quality trait analysis and QTL detection in hexaploid wheat in humid rain-fed agriculture

Humid rain-fed agriculture is a special environment for wheat (Triticum aestivum) culture that tends to negatively affect wheat yield and quality. To identify quality characters of wheat in a humid environment, we conducted quality analysis and quantitative trait loci (QTL) detection in a recombinant inbred line whose parent had a high level of quality for several years. We found that high-quality wheat had less gluten content and lower protein content. Apparently, wheat quality and associated quantity traits were in a dynamic state of equilibrium. We detected 83 QTL for 10 wheat quality traits in this recombinant inbred line population. Nine QTL were detected in both evaluation years; Q.DT.scau-2A, linked to Xwmc522-2A, was detected at the same genetic location in both years. Other QTL for different traits were detected simultaneously in more than one location. Consequently, there appeared to be pleiotropic genes that control wheat quality. Based on previous studies and our research on QTL analysis of grain protein content, we conclude that there must be one or more genes for grain protein content on chromosome 6B, whose expression was little affected by environment. We constructed a consensus map and projected the QTL on it. It was useful for choosing optimal markers for marker-assisted breeding and map-based cloning.

The Foliar and Ear Diseases Dynamic in Autumn Wheat at the ARDS Turda During

The wheat crop is considered the crop with the largest cultivated area in the world. In our country, the area cultivated with wheat, represents about 40% of the area cultivated with cereals. Autumn wheat crops are affected by many diseases that cause significant quantitative and qualitative damages. The production losses varies from year to year, due to weather conditions, applied technology, the sources of inoculum and the cultivated variety. From the foliar and ear diseases encountered in the culture of wheat, powdery mildew ( Erysiphe graminis f.sp. tritici fc. Oidiummonilioides  ), brown staining (Septoria tritici fc. Leptosphaeria tritici ) rust (Puccinia spp.) and fusariosis ( Gibberella spp fc. Fusarium graminearum) occurs with high intensity, especially in the years with high precipitation. In this paper is presented the dynamics of the main wheat diseases in the climatic conditions of the years 2005 - 2011, the ARDS Turda. The experiments were located by random block method in three repetitions. The biological material was represented by four wheat varieties created at ARDS Turda. The observation were made once a week from the end of April - early May until the end of July. The observation were made to 20 plants of each variety. It was noted: the number of siblings, number the leaves, leaf length, leaf width, the attack degree on the plant, specifying at each notation the phenological stage. Following observation in the experimental conditions from ARDS Turda, the frequent diseases were: powdery mildew, brown staining and fusariosa.

Effects of magnetic field on cell dedifferentiation and callus induction derived from embryo culture in bread wheat (Triticum aestivum. L) genotypes

Current study was conducted to investigation of effect of magnetic field on cell dedifferentiation and follow it callus induction derived from mature embryo culture in bread wheat genotypes. For this purpose, a factorial experiment based on completely randomized design was carried out with two wheat genotypes and three level of magnetic field strength (0.0, 8.8 and 17.6 Tesla) in three replications. Callus growth rate (CGR), relative growth rate (RGR), callus relative growth rate (CRGR), percentage of callus water content and percentage of callus induction traits were measured. To sum up, the results showed that differences between wheat genotypes and level of magnetic field strength were significant for some studied treats related to callus induction. The effect of magnetic field levels on CGR (from 0.181 to 0.175), RGR (from 1.442 to 0.655) and CRGR (from 0.052 to 0.022) were decreased with increment of magnetic field intensity.

The auxins centrophenoxine and 2,4-D differ in their effects on non-directly induced chromosome doubling in anther culture of wheat (T. aestivum L.)

Doubled haploid technologies have become key tools for plant breeding. Using these techniques, the speed and efficiency of plant improvement processes can be significantly enhanced. Anther culture-based technologies have the potential to regenerate large numbers of doubled haploid plants without colchicine treatment. In an attempt to elucidate the influence of phytohormones on non-directly induced chromosome doubling, two synthetic auxins, 2,4-D and centrophenoxine, were tested in a wheat anther culture approach. Whereas the induction of androgenic embryo-like structures (ELSs) was efficient for both auxins, we observed a significantly higher frequency of chromosome doubling when using 2,4-D than when using centrophenoxine. When 2,4-D was added to the induction medium, a positive correlation between the size of ELSs and their ploidy level was detected by flow cytometry. The morphological selection of ELSs, a process that was included in routine operations of the method without significantly extending the input of time and effort, facilitates the production of fertile DH plants with a frequency of 60 %. Our findings may contribute to a more efficient production of doubled haploid wheat plants using a colchicine-free anther culture approach.

Special issue: Masked mycotoxins

In 1990, Gareis et al. referred to a zearalenone-glycoside as a ‘masked mycotoxin’ to emphasise the fact that this mycotoxin conjugate was not detected by routine analysis of food or feed, but probably contributed to the total mycotoxin content and subsequent effects. Indeed, pigs fed with mixed feed, artificially contaminated with synthesised zearalenone-4-ß-D-glucopyranoside, excreted zearalenone (ZEA) and α-zearalenol in their faeces and urine, demonstrating release of the aglucone during digestion. Earlier, in 1988, Engelhardt et al. demonstrated that wheat and maize cell cultures were able to transform ZEA to the ß-D-glucopyranoside-conjugate as part of their plant metabolism. It is generally known that plants can reduce the toxicity of phytotoxic compounds by chemical modification. This plant detoxification process includes the conjugation of mycotoxins to polar substances such as sugars, amino acids and sulphate, and subsequent storage of the conjugates in cell vacuoles. In 2002, Schneweis et al. described the natural occurrence of zearalenone-4-glucoside in wheat, while in 2005, Berthiller et al. published the first report on the natural occurrence of deoxynivalenol-3-glucoside (DON-3G) in maize and wheat samples. Since then, research on masked mycotoxins has grown exponentially. Besides plant metabolism, food technological processes also have an impact on the masking mechanism, specifically in cereal-based products in the case of fumonisins (Humpf and Voss, 2004; Dall’Asta et al., 2008) and deoxynivalenol (DON) (Lancova et al., 2008).

Structural changes in chloroplast genome accompanying albinism in anther culture of wheat and triticale

Structural changes in chloroplast genome accompanying albinism in anther culture of wheat and triticale

Structural changes in chloroplast genome accompanying albinism in anther culture of wheat and triticale

The occurrence frequency of structural changes in plastid genome of albino plants from 27 genotypes of Triticum aestivum L. and × Triticosecale Witt. produced by anther culture method was studied. PCR analysis of genome rearrangements in plastids of albino plants has revealed similarity of plasmon changes in the given cereal crops and shown that a number of genes related to photosynthesis exposed to changes in plastid genomes studied. atpB gene, encoding β-subunit of CF1-complex of ATP synthase, was not detected with a high frequency in chlorophyll-deficient plants of wheat and triticale. It is supposed that deletions of photosynthetic genes, especially atpB gene, which take place in evolutionary unstable regions of chloroplast genome with a high frequency, are the major cause of albino plants formation in anther culture of cereals. Genes psbA, psbC, psbD of the photosystem II are amplified practically in all the studied plants of wheat and triticale that verifies the earlier obtained information about conservation of the region including the given genes within the limits of a large singlecopy region of chloroplast DNA.

Susceptibility of common herbaceous crops to Phytophthora cinnamomi and its influence on Quercus root rot in rangelands

The susceptibility to Phytophthora cinnamomi of four crops (wheat, oat, vetch, and yellow lupin) commonly planted in rangeland ecosystems in southern Spain was evaluated. By means of in vitro infection experiments, the presence of the pathogen into the roots of yellow lupin (symptomatic) and vetch (asymptomatic) was observed, but never into wheat and oat roots (asymptomatic). It was also demonstrated that yellow lupin stimulated the production of zoospores of P. cinnamomi. Vetch, wheat and oat did not stimulate zoospore production. Under controlled conditions, only yellow lupin induced an increase in the number of viable chlamydospores in the soil. We concluded that the culture of wheat, oat, and vetch in rangelands did not influence the epidemiology of the Quercus root disease, even when asymptomatically-infected vetch is grown, and these crops can constitute an alternative to the culture of yellow lupin in rangeland ecosystems affected by Quercus root rot.

Phylogenetic analyses and toxigenic profiles of Fusarium equiseti and Fusarium acuminatum isolated from cereals from Southern Europe

Fusarium equiseti and Fusarium acuminatum are toxigenic species that contaminate cereal crops from diverse climatic regions. They are common in Spanish cereals. The information available on their phylogenetics and toxigenic profiles is, however, insufficient to assist risk evaluation. In this work, phylogenetic analyses were performed using partial sequences of the translation elongation factor gene (EF-1α) of F. equiseti and F. acuminatum strains isolated from barley and wheat from Spain and other countries. The Northern and Southern European F. equiseti strains largely separated into two phylogenetically distinct clusters. This suggests the existence of two distinct populations within this species, explaining its presence in these regions of markedly different climate. Production of type A and B trichothecenes by the Spanish strains, examined in wheat cultures using a multitoxin analytical method, indicated that F. equiseti could produce deoxynivalenol and nivalenol and other trichothecenes, at concentrations that might represent a significant risk of toxin contamination for Southern European cereals. F. acuminatum showed low intraspecific genetic variability and 58% of the strains could produce deoxynivalenol at low level. Neither species was found to produce T-2 or HT-2 toxins. The present results provide important phylogenetic and toxigenic information essential for the accurate prediction of toxigenic risk.

Comportamento produtivo do trigo em diferentes métodos e densidades de semeadura

The method and sowing density constitute important factors so that obtained good productivity in wheat culture. The objective this work was evaluated the effects of sowing methods (throwing and in line) and densities (160, 320 and 480 plants m-2) on the performance of the wheat cultivated in no-tillage system. The experiment was conducted in winter period of 2007, with treatments arranged in complete blocks design at random in a factorial arrangement 2 x 3 (sowing methods x sowing density), with four replications. The characteristics were evaluated: tiller number per plant, ears for m2 and spikelets per ear, plant height, ear weight and of one-hundred grains, water tenor in the grains and productivity of the culture. The results were submitted to the variance analysis, the means being the sowing methods compared by Tukey test and the densities for polynomial regression. The plant height was big when the wheat was sowed in line, while, the tiller number per plant, ear weight and of hundred grains and water tenor in the grains was bigger in the throwing sowing. The growth in the sowing density provoked linear reduction of the tiller number of per plant and spikelets per ear, plant height, ear weight and of one-hundred grains. The ears number per m2 increased together with the growth of the density, nevertheless, such an effect did not think about the productivity, and this not was influenced by the respected factors.

High frequency wheat regeneration from leaf tissue explants of regenerated plantlets

The specificities of tissue culture of wheat greatly limit the use of chloroplast transformation technologies in this crop. One limitation in wheat tissue culture is that it is difficult to regenerate plantlets from leaf tissue explants of regenerated plantlets, resulting in difficulty in obtaining homoplastic plants via multiple rounds of antibiotic selection of chloroplast transformants. Thus, a repeated in vitro regeneration system from leaf tissues was studied in this research. Our results showed that 2 mm leaf basal segments of the 4 cm high leaves from regenerated plantlets can give the best callus induction at present study. The best callus induction medium was Murashige and Skoog (MS) basal medium supplemented with 2 mg/L 2,4-dichlorophenoxyacetic acid and 1 mg/L naphthalenacetic acid, which gave a callus induction rate of up to 87.2%. The optimal differentiation medium was MS basal medium supplemented with 10 mg/L silver nitrate and 1 mg/L 2,3,5-triiodobenzoic acid, which gave a regeneration rate up to 33.7% for the wheat lines tested. This is the first report showing that leaf basal segments of in vitro regenerated plantlets can be used for regeneration of wheat. The establishment of a repetitive regeneration system should pave the way for the development of chloroplast transformation and the plant regeneration systems starting from leaf material of in vitro regenerated wheat and other cereal crops.

Inhibition of Fusarium graminearum growth in flour gel cultures by hexane-soluble compounds from oat (Avena sativa L.) flour.

Fusarium head blight, incited by the fungus Fusarium graminearum, primarily affects wheat (Triticum aestivum) and barley (Hordeum vulgarum), while oat (Avena sativa) appears to be more resistant. Although this has generally been attributed to the open panicle of oats, we hypothesized that a chemical component of oats might contribute to this resistance. To test this hypothesis, we created culture media made of wheat, barley, and oat flour gels (6 g of flour in 20 ml of water, gelled by autoclaving) and inoculated these with plugs of F. graminearum from actively growing cultures. Fusarium growth was measured from the diameter of the fungal plaque. Plaque diameter was significantly smaller on oat flour cultures than on wheat or barley cultures after 40 to 80 h of growth. Ergosterol concentration was also significantly lower in oat cultures than in wheat cultures after growth. A hexane extract from oats added to wheat flour also inhibited Fusarium growth, and Fusarium grew better on hexane-defatted oat flour. The growth of Fusarium on oat flour was significantly and negatively affected by the oil concentration in the oat, in a linear relationship. A hexane-soluble chemical in oat flour appears to inhibit Fusarium growth and might contribute to oat's resistance to Fusarium head blight. Oxygenated fatty acids, including hydroxy, dihydroxy, and epoxy fatty acids, were identified in the hexane extracts and are likely candidates for causing the inhibition.

Phosphorus pools and other soil properties in the rhizosphere of wheat and legumes growing in three soils in monoculture or as a mixture of wheat and legume

Phosphorus and nitrogen availability and forms are affected by soil properties as well as by plant species and further modulated by soil microbes. Additionally, close contact of the roots of two plant species may affect concentrations and forms of N and P. The aim of this study was to assess properties related to N and P cycling in the rhizosphere of wheat and legumes grown in monoculture or in wheat/legume mixtures in three soils differing in pH. Faba bean, white lupin and wheat were grown in three soils differing in pH (4.8, 7.5 and 8.8) in monoculture or in mixed culture of wheat and legumes. Rhizosphere soil was collected at flowering and analyzed for P pools by sequential fractionation, available N as well as community structure of bacteria, fungi, ammonia oxidizers, N2-fixers and P mobilizers by polymerase chain reaction (PCR)—denaturing gradient gel electrophoresis (DGGE). Soil type was the major factor determining plant growth, rhizosphere nutrient dynamics and microbial community structure. Among the crop species, only faba bean had a significant effect on nitrification potential activity (PNA) in all three soils with lower activity compared to the unplanted soil. Soil type and plant spieces affected the community composition of ammonia-oxidizing archaea (AOB), ammonia-oxidizing archaea (AOA), N2-fixers (nifH), P mobilizers (ALP gene) and fungi, but not that of bacteria. Among the microbial groups, the AOA and nifH community composition were most strongly affected by crop species, cropping system and soil type, suggesting that these groups are quite sensitive to environmental conditions. All plants depleted some labile as well as non-labile P pools whereas the less labile organic P pools (NaOH extractable P pools, acid extractable P pools) accumulated in the rhizosphere of legumes. The pattern of depletion and accumulation of some P pools differed between monoculture and mixed culture as well as among soils. Plant growth and rhizosphere properties were mainly affected by soil type, but also by crop species whereas cropping system had the least effect. Wheat and the legumes depleted less labile inorganic P pools in some soils whereas less labile organic P pools (NaOH extractable P, acid extractable P) accumulated in the rhizosphere of legumes.

Starch accumulation, activities of key enzyme and gene expression in starch synthesis of wheat endosperm with different starch contents

In order to investigate starch accumulation, and the enzymes activity changes and the expression levels of genes and their relationships among them at different developmental stages of wheat grain. We choose Annong9912 and E28 were used in the study. During starch accumulating rate and grain filling rate, and there were obvious genotype difference between Annong9912 and E28. Whether low or high starch content of starch content, the accumulation courses of amylopectin, amylose and total starch were well fitted to the logistic equation by relating starch contents against DAP. The simulation parameters revealed that the higher contents of amylopectin and amylose resulted from earlier initiating accumulation time and greater accumulation rate. And amylose, amylopectin and total starch accumulation rate of two wheat cultures were significantly and positively correlated with activities of SBE, SSS and GBSS, but amylose accumulation rate of E28 had no correlation with the activities of SBE. In addition, there were significant correlations among activities of SBE, SSS and GBSS in two wheat cultivars. We speculated that these enzymes proteins may have a coordinating action in starch biosynthesis within the amyloplast, operating as functional multiprotein complexes. And expression levels of enzyme genes demonstrated a single-peak curve, and 12-18 DAP reached their peaks and then began to drop, and all had high expression level in earlier stage of endosperm development, but in E28 were higher than in Annong9912. The GBSS-I transcripts on average were expressed over 60 times more than GBSS-II transcript in E28. SBE, SSS, DBE may control starch synthesis at the transcriptional level, and GBSS-I may control starch synthesis at the post transcriptional level. The expression level of DBE on average was lower than SS-1 and SBE-IIa genes, and similar to SS-III and SBE-IIb genes, but higher than GBSS-I and GBSS-II genes.

Competição de trigo com azevém em função de épocas de aplicação e doses de nitrogênio

O potencial de produtividade da cultura do trigo é limitado pela competição exercida pelas plantas daninhas, especialmente pelos recursos luz, água e nutrientes. Em sistemas de produção, a disponibilidade de nitrogênio (N) quase sempre é limitante, influenciando o crescimento da planta mais do que qualquer outro nutriente. O objetivo deste trabalho foi avaliar variáveis morfológicas e componentes da produtividade da cultura do trigo em competição com azevém, em função de épocas de aplicação e doses de N. Os tratamentos foram dispostos em esquema fatorial (2 x 3 x 5), em que o primeiro fator constou de condição de competição (ausência e presença de azevém), o segundo testou épocas de aplicação de N (100% aplicado no início do afilhamento; 50% no início do afilhamento e 50% no início da diferenciação da espiga (IDE); e 100% no IDE) e o terceiro avaliou doses de N (0, 40, 80, 120 e 160 kg ha-1), na forma de ureia aplicada a lanço. As variáveis analisadas foram: área foliar (AF), massa seca da parte aérea (MSPA), número de afilhos (NA), número de colmos (NC), teor de N da cultura (TN), número de grãos por espigueta (NGES), número de grãos por espiga (NGE), número de espiguetas por espiga (NESE), produtividade biológica (PB), massa média de mil grãos (MMG) e produtividade de grãos (P). O azevém compete com o trigo pelo recurso N, reduzindo a AF, MSPA, NA, NC, TN, NGES, NGE, NESE, PB, MMG e P da cultura. A aplicação do N no início do afilhamento ou fracionada, nas duas doses mais elevadas, aumenta a AF e MSPA. O aumento da dose de N incrementa o TN na cultura do trigo, independentemente da presença ou ausência do competidor. A aplicação de N no IDE ou o incremento da dose de N aumenta o NGES e NGE.

Responses of rainwater conservation, precipitation-use efficiency and grain yield of summer maize to a furrow-planting and straw-mulching system in northern China

In the rain-fed areas of northern China, maize ( Zea mays L.) is a main field crop, as it is well adapted to high temperatures and bright sunshine. However, low and variable rainfall and high evapotranspiration rates are common in water-limited environments during the growing season, and often mismatched rainfall events with the critical growth stages, making yield unstable. In this study, the performance of a furrow-planting and straw-mulching system was compared with the conventional flat-planting system in a double-crop culture of winter wheat ( Triticum aestivum L.) and summer maize for two consecutive years (2005–2006 and 2006–2007). The four tested treatments were: conventional flat planting (F), furrow planting between ridges (B), flat planting with wheat straw-mulching (FS), and furrow planting between ridges with wheat-straw mulch (BS). Soil water content and leaf area index (LAI) were measured throughout the growing season each year, and grain yield and precipitation-use efficiency (PUE Y) were determined. On average, ridge tillage combined with furrow planting increased maize yield by 430 kg ha −1 (7.3%) and PUE Y by 10.7% (1.5 kg ha −1 mm −1), compared with the conventional flat planting; furrow planting coupled with straw mulching increased yield by an additional 16.9% and PUE Y by 19.4%, respectively. From jointing to maturity, LAI values of BS were significantly higher than those of F-system (55.6% vs. 26.1% in 2006 and 81.4% vs. 21.7% in 2007). Our data suggest that maize production adopted by furrow planting with straw-covered ridges performed best under seasonal average rainfall below 480 mm, which was associated with better synchronization of seasonal soil water supply and crop needs, leading to improved maize yield and PUE Y.

Microbial ecology of the watery ecosystems of Evros river in North Eastern Greece and its influence upon the cultivated soil ecosystem

The aim of the present study was to evaluate the microbial ecosystem of cultivated soils along the Evros river in NE Greece. Evros river together with its derivative rivers constitute the capital source of life and sustainable development of the area. Along this riverside watery ecosystem systematic agro-cultures were developed such as wheat, corn and vegetable cultures. The evaluation of the ecosystem microbial charge was conducted in both axes which are the watery ecosystem and the riverside cultivated soil area. Considerable discrimination of water quality was observed when considering chemical and microbiological parameters of the Evros river ecosystem. Ardas river possesses a better water quality than Evros and Erythropotamos, which is mainly due to the higher quantities that these two rivers accumulate from industrial, farming and urban residues leading to higher degree of pollution. An increased microbial pollution was recorded in two of the three rivers monitored and a direct relation in microbial and chemical charging between water and cultivated-soil ecosystems was observed. The protection of these ecosystems with appropriate cultivated practices and control of human and animal activities will define the homeostasis of the environmental area.

The application of n-butanol improves embryo and green plant production in anther culture of Australian wheat (Triticum aestivum L.) genotypes

The objective of this study was to improve the production from anther culture of embryos and green plants in Australian spring wheat genotypes by testing new treatments such as n-butanol, as well as other protocol modifications. To date, the use of n-butanol to enhance embryogenesis has only been tested in two European wheat cultivars; this is the first study which demonstrates its application across a range of breeding crosses. A 5-h treatment using 0.1 or 0.2% (v/v) n-butanol following anther pretreatment on a solid mannitol medium significantly improved the production of embryos, green plants and doubled haploids in a range of Australian wheat crosses and varieties. Green plant production increased between 3- and 6-fold in the crosses Yitpi/2*Bumper, Tammarin Rock/2*Bumper and Tammarin Rock/2*Magenta. The addition of calcium (Ca) and macronutrients to the mannitol pretreatment medium also significantly improved the number of embryos and green plants in varieties and crosses, but only when used in combination with n-butanol treatment. A factorial experiment with four varieties and two treatments (n-butanol and Ca/macronutrients) revealed significant interactions between treatments and genotype. In three of the four varieties, the application of n-butanol resulted in significant increases in embryos and green plants with either pretreatment medium although the best results were obtained with Ca and macronutrients in the pretreatment medium, with 200, 193 and 52 green plants per 100 anthers obtained for Bumper, Gladius and Magenta, respectively. In the variety Fortune however, n-butanol treatment did not improve embryo or green plant production unless it was combined with Ca and macronutrients in the pretreatment medium and then there were dramatic improvements; from 0 to 27 green plants per 100 anthers.