Isolation and characterisation of microsatellites from hexaploid bread wheat

 The development of large panels of simple-to-analyse genetic markers for tagging agronomically important genes and diversity studies in hexaploid bread wheat is an important goal in applied cereal genetic research. We have isolated and sequenced over 200 clones containing microsatellites from the wheat genome and have tested 153 primer pairs for genetic polymorphism using a panel of ten wheat varieties, including the parents of our main mapping cross. A subset comprising 49 primer pairs detects 76 loci, of which 74 can be unequivocably allocated to one of the wheat chromosomes. A relatively low frequency of the loci detected are from the D genome, and these loci show less polymorphism than those from the A and B genomes. Generally, the microsatellites show high levels of genetic polymorphism and an average of 3.5 alleles per locus with an average polymorphism information content (PIC), value of 0.51. The observed levels of polymorphism are positively correlated with the length of the microsatellite repeats. A high proportion, approximately two-thirds, of primer pairs designed to detect simple sequence repeat (SSR) variation in wheat do not generate the expected amplification products and, more significantly, often generate unresolvable PCR products. In general, our results agree closely with those obtained from other recent studies using microsatellites in plants. Received: 19 March 1996 / Accepted: 28 June 1996     

Separation of bioactive biflavonoids from Rheedia gardneriana using chitosan modified with benzaldehyde

This paper shows the influence of benzenic groups on the chitosan surface for the separation of bioactive biflavonoids from Rheedia gardneriana leaves The yield of the biflavonoids using chitin modified with benzaldehyde (CH-Bz) as adsorbent in column chromatography was higher than that achieved with silica gel and chitosan. The presence of benzenic groups decreases the polarity of chitosan and consequently the interaction of hydrogen bonding between phenolic hydroxyl (OH) of biflavonoids and amine groups of the adsorbent. Therefore, the separation of these compounds appears to be the result of hydrophobicity and pi-pi interaction among electrons from the aromatic ring in sorbent and biflavonoid molecules.     

Separation of flavonoids from Aleurites moluccana leaves using chitosan modified with heptaldehyde

Heptaldehyde-modified chitosan (heptyl-chitosan, CH-Hp) was investigated as adsorbent for chromatograhic separation of the flavonoids from A. moluccana. The amount of 2"-O-rhamnosylswertisin isolated (30.0 mg) was approx. twice as high as swertisin (17.5 mg). The improved surface hydrophobicity effected by the heptyl groups promoted the separation of flavonoids. From the results obtained, CH-Hp seems to be more suitable for separation of glycosylated flavonoids than other flavonoids. Thus, modified chitosan described here can be used for hydrophobic interaction chromatography as sucessfully illustrated with flavonoids.     

Transfer of the 1BL/1RS wheat-rye-translocation from hexaploid bread wheat to tetraploid durum wheat

Summary The present study describes a cytological stable alien chromosome translocation in tetraploid durum wheat. By crossing the hexaploid 1BL/1RS wheat-rye translocation line Veery to the tetraploid durum wheat cultivar Cando it was possible to select a 28 chromosomic strain homozygous for the 1BL/1RS translocation. The disease resistance potential of the short arm of rye chromosome 1R, which has been widely introduced in many hexaploid bread wheat cultivars could be now also used for the improvement of durum wheat.     

Making bread with sourdough improves iron bioavailability from reconstituted fortified wheat flour in mice

This study aimed to evaluate the effect of a diet prepared with traditional sourdough (TS) on iron status. Levels of blood hemoglobin (Hb), Hematocrite (Ht), serum ferritin and serum iron as well as excreted iron were determined in three groups of mice fed with: TS bread (TS group), baking yeast bread (BY group) or bread with no starters (control group), respectively. The results show that the levels of Hb, Ht, ferritin and iron were significantly higher in the TS compared to the BY and control groups. Also a significant decrease in the excreted iron levels was observed in the mice fed with TS compared to the others dietary groups.

In conclusion, the study results indicate an improvement of iron status indicators in mice when they were fed sourdough bread as compared to baking yeast bread and bread with no starters.     

Identification of genetic loci associated with ear-emergence in bread wheat

A doubled haploid population constructed from a cross between the South Australian wheat cultivars ‘Trident’ and ‘Molineux’ was grown under winter field conditions, under field conditions over summer and under artificial light both with and without vernalisation. The duration from planting to ear-emergence was recorded and QTL associated with heading date were detected using a previously constructed genetic linkage map. Associations were shown with chromosomal regions syntenous to previously identified photoperiod (Ppd-B1) and vernalisation (Vrn-A1) sensitive loci. Additional QTL associated with time to heading were also identified on chromosomes 1A, 2A, 2B, 6D, 7A and 7B. Comparisons between the genetic associations observed under the different growing conditions allowed the majority of these loci to be classified as having either photoperiod-sensitive, vernalisation-sensitive or earliness per se actions. The identification of a photoperiod-sensitive QTL on chromosome 1A provides evidence for a wheat gene possibly homoeologous to Ppd-H2 previously identified on chromosome 1H of barley. The occurrence of a putative major gene for photoperiod sensitivity observed on chromosome 7A is presented. The combined additive effects at these loci accounted for more than half the phenotypic variance in the duration from planting to ear-emergence in this population. The possible role of these loci on the adaptation of wheat in Australia is discussed.     

Integration of dinucleotide microsatellites from hexaploid bread wheat into a genetic linkage map of durum wheat

 Seventy nine microsatellite markers from hexaploid bread wheat (T. aestivum L.) were integrated into a genetic linkage map of durum wheat (T. turgidum ssp. durum (Desf.) Huns.) created by RFLP segregation data from a population of 65 recombinant inbred lines. The results indicate a relatively even distribution of microsatellite loci and demonstrate that microsatellite markers from hexaploid wheat provide an excellent source of molecular markers for use in the genetics and breeding of durum wheat. Received: 16 July 1998 / Accepted: 13 October 1998     

Molecular-genetic analysis of the breeding bread wheat lines with starch of amylopectin type

PCR-analysis with primers to Wx-loci of T. aestivum L. genome lets us run the controlled selection of genotypes with null-alleles of Wx-genes while creating bread wheat lines with amylopectin type of starch. Microsatellite analysis of selected individual plants which were the carries of three null-alleles of Wx-genes (Wx-A1b, Wx-B1b, Wx-D1b) and the cluster analysis in the complex allowed us to pick out four genotypes, which were very close related genetically to the parent form variety Kuyalnik.     

Characteristics of Egyptian boza and a fermentable yeast strain isolated from the wheat bread

A sample of indigenous beer, boza was collected at Cairo, Egypt and analysed. Boza was an off-white porridge-like slurry containing 3.8% (v/v) ethanol. Volatile esters and higher alcohols such as ethyl acetate and isoamyl alcohol were detected in the boza by gas chromatography. The pH of the boza was 3.7. Organoleptically, this alcoholic beverage had an estery flavour and a sour taste. A fermentable yeast strain EG1 was isolated from the material wheat bread and identified, and was considered to resemble Candida krusei. The rice sake made with the yeast strain C. krusei EG1 at 30 °C contained 11.7% ethanol, 74.1 mg/l ethyl acetate and its pH value was 4.2.     

Modeling and phylogeny analysis of bread wheat MnSOD

Superoxide dismutase (SOD) acts as first line of defense against oxidative and genetic stress. Manganese superoxide dismutase (MnSOD), found in mitochondria or peroxisomes, contains Mn(III) at the active site. Therefore, it is of interest to study MnSOD from bread wheat (a grain crop). However, a structure model is not yet solved for bread wheat MnSOD. Hence, we describe the structure model of bread wheat MnSOD developed using homology model. The model provides molecular insight to metal binding molecular function towards the understanding of oxidative stress resistance in plants. The distinction of bread wheat (a monocot) MnSOD from dicots is also shown using phylogenetic analysis.     

Marking the Vrd1 gene in the bread winter wheat

PCR analysis was used to create DNA markers to the Vrd1 gene. DNA of almost isogenic lines with respect to Vrd genes of the cultivars Mironovskaya 808 and Erytrospermum 604 was used. It was shown that in the monogenic Vrd1 dominant genotypes the product of amplification (280 b.p.) is absent in comparison with the DNA of the vrd recessive and monogenic Vrd2 dominant genotypes. The linkage of the marker with the Vrd1 gene has been determined using DNA analysis of plant population obtained as a result of crossing of Erytrospermum 604 (vrd recessive) and Triple Dirk C (Vrd1vrd2). F2 population segragated in two groups on the character of 280 b.p. amplification product "presence/absence". The segregation significantly coincided to the theoretical one (by ?2 test) with 1:3 expectation. The revealed molecular marker identified homozygous dominant Vrd1 plants only. The DNA-marker to Vrd1 gene is nulle-allelic 280(-).     

Microsatellite-based molecular diversity of bread wheat germplasm and association mapping of wheat resistance to the Russian wheat aphid

Genetic diversity of a set of 71 wheat accessions, including 53 biotype 2 Russian wheat aphid (RWA2)-resistant landraces and 18 RWA2 susceptible accessions, was assessed by examining molecular variation at multiple microsatellite (SSR) loci. Fifty-one wheat SSR primer pairs were used, 81 SSR loci were determined, and 545 SSR alleles were detected. These SSR loci covered all the three genomes, 21 chromosomes, and at least 41 of the 42 chromosome arms. Diversity values averaged over SSR loci were high with mean number of SSR alleles/locus = 6.7, mean Shannon’s index (H) = 1.291, and mean Nei’s gene diversity (He) = 0.609. The three wheat genomes ranked as A > D > B and the homoeologous groups ranked as 7 > 3  > 1 > 2 > 6 > 5 > 4 based on the number of alleles per locus. Xgwm136 on chromosome arm 1AS is the most polymorphic SSR locus with the largest number of observed and effective alleles and the highest H and He. Among all 2485 pairs of wheat accessions, genetic distance (GD) ranged from 0.054 to 1.933 and averaged 0.9832. A dendrogram based on GD matrix showed that all the wheat accessions could be grouped into distinct clusters. Most of the susceptible cultivars (13/18) were clustered into groups that contains all or mostly susceptible accessions. Most of the U.S. cultivars belong to a group that is distinguishable from all the different RWA2 resistant groups. Diversity analysis was also conducted separately for subgroups containing 53 RWA2-resistant accessions and 18 RWA2-susceptible accessions. Association mapping revealed 28 SSR loci significantly associated with leaf chlorosis, and 8 with leaf rolling. New chromosome regions associated with RWA2 resistance were detected, and indicated existence of new RWA resistance genes located on chromosomes of all other homoeologous groups in addition to the groups 1 and 7 in bread wheat. This information is helpful for development of mapping populations for RWA2 resistance genes from different phylogenetic groups, and for wise utilization of the RWA-resistant germplasm in wheat breeding programs.     

Characterisation of polymorphic microsatellite markers from Aegilops tauschii and transferability to the D-genome of bread wheat

Microsatellites were isolated from a Aegilops tauschii (the D-genome donor of bread wheat) library enriched for various motifs. Primers generated from the flanking region of the microsatellites were used successfully to amplify the corresponding loci in the D genome of bread wheat. Additional amplification sometimes also occurred from the A and B genomes. The majority of the microsatellites contained (GA)(n) and (GT)(n) motifs. GA and GT repeats appeared to be both more abundant in this library and more polymorphic than other types of repeats. The allele number for both types of dinucleotide repeats fitted a Poisson distribution. Deviance analysis showed that GA and GT were more polymorphic than other motifs in bread wheat. Within each motif type (di-, tri- and tetra-nucleotide repeats), repeat number has no influence on polymorphism. The microsatellites were mapped using the Triticum aestivum Courtot x Chinese Spring mapping population. A total of 100 markers was developed on this intraspecific map, mainly on the D genome. For polyploid species, isolation of microsatellites from an ancestral diploid donor seems to be an efficient way of developing markers for the corresponding genome in the polyploid plant.     

Diversity of wheat anti-microbial peptides

From seeds of Triticum kiharae Dorof. et Migusch., 24 novel anti-microbial peptides were isolated and characterized by a combination of three-step HPLC (affinity, size-exclusion and reversed-phase) with matrix-assisted laser-desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry and Edman degradation. Based on sequence similarity and cysteine motifs, partially sequenced peptides were assigned to 7 families: defensins, thionins, lipid-transfer proteins, hevein-like peptides, knottin-like peptides, glycine-rich peptides, and MBP-1 homologs. A novel subfamily of defensins consisting of 6 peptides and a new family of glycine-rich (8 peptides with different repeat motifs) were identified. Three 6-cysteine knottin-like peptides represented by N- and C-terminally truncated variants revealed no sequence homology to any known plant anti-microbial peptides. A new 8-cysteine hevein-like peptide and three 4-cysteine peptides homologous to MBP-1 from maize were isolated. This is the first communication on the occurrence of nearly all families of plant anti-microbial peptides in a single species.     

Mapping and validation of quantitative trait loci associated with wheat yellow mosaic bymovirus resistance in bread wheat

Wheat yellow mosaic (WYM) caused by wheat yellow mosaic bymovirus (WYMV) has been growing as one of the most serious diseases affecting wheat production in China. In this study, the association of quantitative trait loci (QTLs) governing WYMV resistance with molecular markers was established using 164 recombinant inbred lines (RILs) derived from 'Xifeng Wheat' (highly resistant)?×?'Zhen 9523' (highly susceptible). Phenotypic data of WYMV resistance of the RILs were collected from 4-year, two-location replicated field trials. A molecular marker-based linkage map, which was comprised of 273 non-redundant loci and represented all the 21 wheat chromosomes, was constructed with the JoinMap 4.0 software. Using the Windows QTL Cartographer V2.5 software, three QTLs associated with WYMV resistance, QYm.njau-3B.1, QYm.njau-5A.1 and QYm.njau-7B.1, were detected on chromosomes 3BS, 5AL, and 7BS, respectively. The favorable allele effects were all contributed by 'Xifeng Wheat'. Among the three QTLs, QYm.njau-3B.1 and QYm.njau-5A.1 were detected in all the four trials and the overall mean, and could explain 3.3-10.2% and 25.9-53.7% of the phenotypic variation, respectively, while QYm.njau-7B.1 was detected in one trial and the overall mean and explained 4.9 and 3.3% of the phenotypic variation, respectively. A large portion of the variability for WYMV response was explained by a major QTL, QYm.njau-5A.1. The relationship of the molecular markers linked with QYm.njau-5A.1 and the WYMV resistance was further validated using a secondary F(2) population. The results showed that three markers, i.e., Xwmc415.1, CINAU152, and CINAU153, were closely linked to QYm.njau-5A.1 with the genetic distances of 0.0, 0.0, and 0.1?cM, respectively, indicating they should be useful in marker-assisted selection (MAS) wheat breeding for WYMV resistance. A panel of germplasm collection consisting of 46 wheat varieties with known WYMV response phenotypes was further used to validate the presence and effects of QYm.njau-5A.1 and the above three markers. It was found that QYm.njau-5A.1 was present in 12 of the 34 WYMV-resistant varieties.     

Enhanced ethanol and chitosan production from wheat straw by Mucor indicus with minimal nutrient consumption

Recently, Mucor indicus was introduced as a promising ethanol producing microorganism for fermentation of lignocellulosic hydrolysates, showing a number of advantages over Saccharomyces cerevisiae. However, high nutrient requirement is the main drawback of the fungus in efficient ethanol production from lignocelluloses. In this study, application of fungal extract as a potential nutrient source replacing all required nutrients in fermentation of wheat straw by M. indicus was investigated. Wheat straw was pretreated with N-methylmorpholine-N-oxide (NMMO) at 120 °C for 1–5 h prior to enzymatic hydrolysis. Hydrolysis yield was improved at least by 6-fold for 3 h pretreated straw compared with that of untreated one. A fungal extract was produced by autolysis of M. indicus biomass, an unavoidable byproduct of fermentation. Maximum free amino nitrogen (2.04 g/L), phosphorus (1.50 g/L), and total nitrogen (4.47 g/L) as well as potassium, magnesium, and calcium in the fungal extract were obtained by autolysis of the biomass at 50 °C and pH 5.0. The fungal extract as a nutrient-rich supplement substituted yeast extract and all other required minerals in fermentation and enhanced the ethanol yield up to 92.1% of the theoretical yield. Besides, appreciate amounts of chitosan were produced as another valuable product of the autolysis.     

Comparative performance of bread wheat and hexaploid triticale cytoplasms

Summary Thirteen wheat-like advanced-generation triticale x wheat derivatives, having tetraploid wheat cytoplasm from triticale, were reciprocally crossed with three improved bread wheats, and the resulting F₁s were evaluated for determining the comparative performance of the bread wheat and triticale cytoplasms for different traits. Significant reciprocal differences in the mean performance were observed for days to heading, days to maturity, spikes/plant, flag-leaf area, peduncle length, plant height, spike length, grains/spike, 1,000-grain weight, grain yield and grain protein content, and most of them were in favour of hexaploid wheat cytoplasm. However, this superiority of the hexaploid cytoplasm was not universal for a particular trait, implying that the differences in the performance of the evaluated reciprocal crosses depended not solely on the cytoplasmic background, but also on the interplay of the specific genotype with the cytoplasm.