Chalcone synthase expression and pigments deposition in wheat with purple and blue colored caryopsis

Red and white caryopses are typical in common wheat. Genotypes with purple and blue caryopses are also described. This coloring is caused by anthocyanins which deposit in the pericarp (purple) or aleurone layer (blue). The anthocyanins biosynthetic pathway is well described. The key enzyme is chalcone synthase (CHS). It catalyzes the first step. We observed the deposition of anthocyanins in the pericarp and aleurone layer, the expression of a chalcone synthase gene and the amount of two anthocyanins - cyanidin-3-glycoside (pericarp of purple caryopses), and delphinidin-3-glycoside (aleurone layer of blue caryopses) during caryopsis development. Purple pigment deposition was not homogeneous and/or uniform. At first, small isolated spots of purple color were formed and thereafter they expanded. In blue caryopses, however, the coloring process was more homogeneous. The expression of chalcone synthase mRNA occurred five days before pigment deposition and finished earlier than expected. Amounts of cyanidin-3-glycoside and delphinidin-3-glycoside increased in a similar manner. Amounts of these fell at the end of caryopses development probably due to formation of more complex substances.     

Starch granules size distribution in superior and inferior grains of wheat is related to enzyme activities and their gene expressions during grain filling

Mature wheat endosperm contains A-, B-, C-type starch granules, and each class has unique physiochemical properties which determine the quality of starch. The dynamics of the starch granule size distribution, activities of starch synthases and expression of starch synthase encoding genes were studied in superior and inferior grains during grain filling. Compared with inferior grains, superior grains showed higher grain weight, contents of starch, amylose and amylopectin. The formation of A-, B-, C-type starch granules initiated at 4, 8, 20 DAF, respectively, and was well consistent with the temporally change patterns of starch synthase activities and relative gene expression levels. For instance, activities of soluble and granule-bound starch synthases (designated SSS and GBSS) peaked at 20 and 24 DAF. Genes encoding isoforms of starch synthases expressed at different grain filling periods. In addition, SS I was generally expressed over the grain filling stage; the SS II and SS III were expressed over the early and mid grain filling stage, and the GBSS I was expressed during the mid to late grain filling stage. In addition, the time-course changes in activities of starch synthases and expression of starch synthase encoding genes explained well the dynamics of the starch granule size distribution.     

Analysis of starch swelling power in Australian breeding lines of hexaploid wheat (Triticum aestivum L.)

Starch is the major component of wheat (Triticum aestivum L.) grain and is composed of two large glucan molecules, amylose and amylopectin. The ratio between the two polymers types influences the water absorbing properties of starch upon heating, and thus affects the end-use of grain and purified starch. In this study, we evaluated the starch swelling power (SSP) values in seven wheat populations developed from crosses involving low-SSP lines. Analysis of starch produced by the F₂ generation plants showed that the largest SSP variation (11.4–16.2) and lowest SSP mean (13.9) was obtained for a population derived from doubled haploid lines SM1028 (SSP = 14.5) and VK306 (SSP = 13.6). The population of 360 lines was advanced by single seed descent to the following generations for further studies. Starch analysis of grain produced by F₄ generation lines in two field locations during 2006 and in a greenhouse environment during 2005 showed that SSP values were relatively stably inherited. The average broad-sense heritability was 73% and significant (P < 0.001) genotype × genotype and genotype × environment interactions were seen. Starches with the highest and lowest SSP values were inversely related to amylose concentration determined by high pressure liquid chromatography (HPLC)–size exclusion chromatography (SEC) of debranched starch. Developed lines with the lowest SSP values surpassed 40% in apparent amylose concentration. The study illustrates that screening for SSP in early generations can be used to develop wheat lines with desired starch swelling characteristics.     

Are seeding densities an opportunity to increase grain yield of winter wheat in a living mulch of white clover?

Optimum plant densities are a key to maximise yields in most crops. However, such information is often lacking for more environmentally sound cropping systems, such as living mulches (LM) for small grains. In 2004 and 2005, three trials were conducted in the Swiss Midlands on fields managed in accordance with the Swiss organic farming guidelines. The objective of the study was to determine whether seeding density of winter wheat (Triticum aestivum L.) is a relevant factor for determining grain yield in a white clover (Trifolium repens L.) living mulch. The winter wheat cv. Titlis was directly sown in wide spaced rows (0.375 m) at densities of 300 (LM300), 450 (LM450) or 600 (LM600) viable grains m⁻² in a white clover living mulch established at a seeding rate of 15 kg ha⁻¹. A bare soil control treatment with a wheat density of 450 viable grains m⁻² (BS450) was also included in the trials. Mean grain yields of LM300, LM450, and LM600 never reached the values observed in BS450. This was mainly due to a lower ear density, which, nevertheless, increased linearly with the seeding density within the living mulch in all trials, but the rate of increase depended on the environment. The decrease of the grain weight brought about by the increasing seeding density had only a marginal impact on the grain yield, which was increased from 1.31, 1.98, and 4.09 Mg ha⁻¹ (LM300) to 1.97, 2.64, and 4.75 Mg ha⁻¹ (LM600) for each of the three trials in the study. Significantly higher protein contents were observed for LM300 compared to the higher densities in the living mulch and to BS450. Our research showed that an increase of the seeding density is an effective mean to increase the grain yield in living mulch systems with white clover. However, it is likely that the control of the living mulch to reduce competition with the main crop is a more relevant factor.     

Genetic characterization of kernel polyphenol oxidases in wheat and related species

Polyphenol oxidase (PPO) activity causes undesirable darkening of raw Asian noodles and other wheat products. In this study we investigate the genetic origins and diversity of wheat kernel PPO. PPO was characterized via activity assays, antigenic staining, and Southern blots in Triticum aestivum, Triticum dicoccoides, Triticum durum, Triticum dicoccum, Triticum monococcum, Triticum urartu, Aegilops speltoides, and Aegilops tauschii. Among these species, PPO activity was well-correlated with antigenic staining intensity toward a wheat kernel-type PPO antibody. High PPO activity was observed in all three T. monococcum accessions (A^m genome), one Ae. speltoides accession, one T. durum accession, and two hexaploid wheat cultivars. Southern blots suggested the presence of two or more kernel-type PPO genes in diploid progenitors of the hexaploid A, B, and D genomes. Whole-kernel PPO activity was evaluated in disomic substitution lines derived from three T. dicoccoides accessions in the background of T. durum ‘Langdon’. PPO activity was primarily associated with chromosome 2A and to a much lower degree with chromosome 2B. DNA sequence comparisons showed that the intron associated with the high PPO allele on chromosome 2AL of hexaploid wheat had 94% nucleotide identity with the homeologous intron found in T. monococcum, a species with high kernel PPO activity. This implies that the ancestral PPO allele on the A genome is one of the high activity, and the low PPO allele found in hexaploid wheat represents a relatively recent genetic alteration. Results confirm the presence of multiple kernel-type PPO genes in the diploid and tetraploid progenitors and relatives of hexaploid wheat. However, it is likely that relatively few of the many kernel-type PPO genes present in wheat contribute substantially to kernel PPO activity. A single genetic locus on homeologous group 2 chromosomes may be the primary cause of high PPO activity in wheat kernels.     

Detection of mutations in the 7A allele of wheat (Triticum aestivum) granule-bound starch synthase (Wx-7A) with a monoclonal antibody produced by targeted peptide immunisation

A peptide sequence unique for the granule-bound starch synthase (GBSS1) of hexaploid wheat (Triticum aestivum) was identified by sequence alignment of the three isoforms. Multiple antigenic peptides (MAP) of different length (16 and 20 AA) were synthesised against the selected sequence. The assessment of the immune response in mice against differently presented forms (resin-bound, free) and two types of adjuvants indicated that the free MAP with the longer peptide is more immunogenic. A 20AA MAP that elicited a strong immune response was used to produce a monoclonal antibody for the 7A isoform of GBSS 1. Epitope mapping of the selected Mab (F5-1F2) revealed that a 12mer partial sequence of the immunising peptide was specifically detected. Based on this Mab, a simple high throughput ELISA was developed that allows the quick identification of wheat lines carrying the 7A allele of GBSS 1 with minute amounts of sample.     

Waxy endosperm accompanies increased fat and saccharide contents in bread wheat (Triticum aestivum L.) grain

The contents of fat, starch, pentosan, fructan, β-glucan and several mono- and oligosaccharides in grain were evaluated to find out the possible effects of the Wx-D1 gene of bread wheat using two sets of near-isogenic waxy and non-waxy lines and two low-amylose mutant lines with a common genetic background of Kanto 107. These materials have two non-functional Wx-A1b and Wx-B1b alleles in common. Waxy near-isogenic lines with a non-functional Wx-D1d allele showed consistently increased contents of fat, total fructan, β-glucan, glucose, fructose, sucrose, 1-kestose, 6-kestose, neokestose, nystose and bifurcose compared with non-waxy lines with a functional Wx-D1a allele throughout three growing/harvest seasons. Starch and total pentosan contents were inconsistently influenced by the allelic status of the Wx-D1 locus, while water-soluble pentosan and raffinose contents were not affected. The compositional changes of a low-amylose mutant line with an almost non-functional Wx-D1f allele were closely similar to those of waxy near-isogenic lines, while significantly different changes were barely observed in another low-amylose mutant line with a partly functional Wx-D1g allele in two seasons. These results showed that the Wx-D1 gene has pleiotropic effects on the fat and saccharide contents of bread wheat grain.     

A MALDI-TOF based analysis of high molecular weight glutenin subunits for wheat breeding

High molecular weight glutenin subunits play an important role in determining wheat dough quality as they confer visco-elastic properties to the dough required for mixing and baking performance. In this work, a collection of 103 genotypes of common wheat from 12 countries was used to analyse the composition of HMW-GS by SDS-PAGE and MALDI-TOF-MS. Results indicated that MALDI-TOF technology is suitable for analyzing most HMW-GS alleles. The allelic diversity at Glu-B1 locus include subunits 6+8b^*, 7, 7+8, 7+8a^*, 7b^*+8, 7^OE, 7^OE+8, 7^OE+8a^*, 7^OE+8b^*, 7+9, 13+16, 14+15, 17+18 and 20. The rapid identification of HMW-GS capability of MALDI-TOF-MS is discussed in relation to its value for screening lines in wheat breeding programs, especially in discriminating subunits 7^OE, 8a^* and 8b^* associated with superior quality. A new glutenin subunit 7b^*+8 was found in Japanese germplasm Eshimashinriki.     

In vitro pullulanase activity of wheat (Triticum aestivum L.) limit-dextrinase type starch debranching enzyme is modulated by redox conditions

Expression of a limit-dextrinase (LD) type starch debranching enzyme (EC 3.2.1.41) was observed in developing wheat (Triticum aestivum L.) endosperm and germinating grains, indicating a role for the enzyme in both biosynthesis and degradation of starch. A full-length cDNA, TaLD1, encoding LD in wheat developing kernels was isolated and predicted to encode a 98.6 kDa mature protein active in amyloplasts. Isolated cDNA was expressed in Escherichia coli to produce a recombinant His-tagged LD, which mainly accumulated in inclusion bodies as an inactive enzyme. Extraction of His-tagged LD from the inclusion bodies followed by dialysis under thiol/disulphide redox conditions allowed partial refolding of the protein and detection of pullulanase specific activities by zymogram analysis and enzyme assays. Several active conformations were demonstrated by the recombinant TaLD1 and pullulanase activity could be modulated by redox conditions in vitro. The results suggest that cellular redox conditions may regulate the level of LD activity in wheat tissues.     

Genetic gains in grain yield, net photosynthesis and stomatal conductance achieved in Henan Province of China between 1981 and 2008

Knowledge of the changes in agronomic and photosynthetic traits associated with genetic gains in grain yield potential is essential for an improved understanding of yield-limiting factors and for determining future breeding strategies. The objectives of this study were to identify agronomic and photosynthetic traits associated with genetic gains in grain yield of facultative wheat (Triticum aestivum L.) between 1981 and 2008 in Henan Province, the most important wheat producing area in China. During the 2006-2007 and 2007-2008 crop seasons, a yield potential trial comprising 18 leading and new cultivars released between 1981 and 2008 was conducted at two locations, using a completely randomised block design of three replicates. Results showed that average annual genetic gain in grain yield was 0.60% or 51.30 kg ha⁻¹ yr⁻¹, and the significant genetic improvement in grain yield was directly attributed to increased thousand grain weight which also contributed to the significant increase in harvest index. The genetic gains in rates of net photosynthesis at 10, 20 and 30 days after anthesis were 1.10% (R² = 0.46, P < 0.01), 0.68% (R² = 0.31, P < 0.05) and 6.77% (R² = 0.34, P < 0.05), respectively. The rates of net photosynthesis at 10 (r = 0.58, P < 0.05), 20 (r = 0.59, P < 0.05) and 30 (r = 0.65, P < 0.01) days after anthesis were closely and positively correlated with grain yield. A slight decrease in leaf temperature and an increase in stomatal conductance after anthesis were also observed. Grain yield was closely and positively associated with stomatal conductance (r = 0.69, P < 0.01) and transpiration rate (r = 0.63, P < 0.01) at 30 days after anthesis. Therefore, improvement of those traits was the likely basis of increasing grain yield in Henan Province between 1981 and 2008. The genetic improvement in yield was primarily attributed to the utilization of two elite parents Yumai 2 and Zhou 8425B. The future challenge of wheat breeding in this region is to maintain the genetic gain in grain yield and to improve grain quality, without increasing inputs for the wheat-maize double cropping system.     

Effect of nitrogen application rate on content of glutenin macropolymer and high molecular weight glutenin subunits in grains of two winter wheat cultivars

Two winter wheat (Triticum aestivum L.) cultivars differing in grain protein content were selected to study the effect of N application rate on changes in contents of glutenin macropolymer (GMP) and high molecular weight glutenin subunits (HMW-GS) during grain filling. Contents of GMP and HMW-GS were much higher in the high GPC cultivar, Xuzhou 26, than those in low GPC cultivar, Ningmai 9. N increased contents of GMP and HMW-GS in Xuzhou 26 with N rate between 0 and 300 kg ha⁻¹, while at the very high N rate of 300 kg ha⁻¹ the contents of GMP and HMW-GS in Ningmai 9 decreased. The high contents of GMP and HMW-GS at maturity were closely related to the rapid increase in contents of GMP and HMW-GS during the initial period of their synthesis. HMW-GS and GMP content were closely correlated. The total HMW-GS content was important in determining GMP content than the content of any HMW-GS pair or any individual HMW-GS present in the selected cultivars. The pattern of response of GMP content to N application rate was closely related to the regulatory effect of N on HMW-GS synthesis.     

Analyses of albumins, globulins and amphiphilic proteins by proteomic approach give new insights on waxy wheat starch metabolism

Starch is composed of two types of glucose polymers: amylose and amylopectin. The Waxy (Wx) locus controls amylose synthesis in the wheat kernel. Hexaploid wheat has three Wx loci located on chromosomes 7A (Wx-A1), 4A (Wx-B1), and 7D (Wx-D1). Eight near isogenic lines (NILs) of Triticum aestivum cv. Tremie with one, two or three Wx null alleles were used. The albumin–globulin fraction, and amphiphilic proteins were separated using 2-dimensional electrophoresis (2DE) allowing the changes in the waxy kernel to be identified. Albumin–globulin fraction showed overexpression of sucrose synthases in the waxy NILs compared to the normal form of Tremie and a decrease in many proteins related to stress and defence metabolism such as serpins. A subunit of ADP-glucose pyrophosphorylase (AGPase), which is known to play a major role in starch synthesis, was also shown to be down regulated in the waxy NILs. Amphiphilic proteins confirmed the observations made on the albumin–globulin fraction with a decrease in a stress-related protein. These different regulations linked to observations made on wheat kernel (thousand kernel weight (TKW), protein amount per grain, size and distribution of the starch granules) led to formulation of the hypothesis that waxy endosperm does not reach maturity of the wild-type endosperm.     

Effects of transition season management on soil N dynamics and system N balances in rice–wheat rotations of Nepal

In the low-input rice–wheat production systems of Nepal, the N nutrition of both crops is largely based on the supply from soil pools. Declining yield trends call for management interventions aiming at the avoidance of native soil N losses. A field study was conducted at two sites in the lowland and the upper mid-hills of Nepal with contrasting temperature regimes and durations of the dry-to-wet season transition period between the harvest of wheat and the transplanting of lowland rice. Technical options included the return of the straw of the preceding wheat crop, the cultivation of short-cycled crops during the transition season, and combinations of both. Dynamics of soil N_min, nitrate leaching, nitrous oxide emissions, and crop N uptake were studied throughout the year between 2004 and 2005 and partial N balances of the cropping systems were established. In the traditional system (bare fallow between wheat and rice) a large accumulation of soil nitrate N and its subsequent disappearance upon soil saturation occurred during the transition season. This nitrate loss was associated with nitrate leaching (6.3 and 12.8 kg ha⁻¹ at the low and high altitude sites, respectively) and peaks of nitrous oxide emissions (120 and 480 mg m⁻² h⁻¹ at the low and high altitude sites, respectively). Incorporation of wheat straw at 3 Mg ha⁻¹ and/or cultivation of a nitrate catch crop during the transition season significantly reduced the build up of soil nitrate and subsequent N losses at the low altitude site. At the high altitude site, cumulative grain yields increased from 2.35 Mg ha⁻¹ with bare fallow during the transition season to 3.44 Mg ha⁻¹ when wheat straw was incorporated. At the low altitude site, the cumulative yield significantly increased from 2.85 Mg ha⁻¹ (bare fallow) to between 3.63 and 6.63 Mg ha⁻¹, depending on the transition season option applied. Irrespective of the site and the land use option applied during the transition season, systems N balances remained largely negative, ranging from −37 to −84 kg N ha⁻¹. We conclude that despite reduced N losses and increased grain yields the proposed options need to be complemented with additional N inputs to sustain long-term productivity.     

Partial purification and characterization of polyphenoloxidase from durum wheat (Triticum durum L.)

A bright yellow color of pasta is an important qualitative trait for the durum wheat industry. Final color is the result of the balance between yellow and brown components in semolina. Polyphenoloxidase (PPO) is implicated as playing a significant role in darkening. This study aimed to characterize PPO activity of durum wheats. PPO was extracted and partially purified by ion-exchange chromatography on a column packed with diethyaminoethyl cellulose (DEAE). This procedure led to 26.33-fold purification with 24.7% recovery. The optimum temperature and pH of PPO were found to be 40 °C and 6.5, respectively. Heat stability of durum wheat PPO decreased as the temperatures increased from 30 to 80 °C. The z-value was calculated as 23.4 °C. It increased to 26.3 and 48.4 °C in the presence of 40% sucrose and 1 M NaCl, respectively. Durum wheat PPO was shown to use several phenolic compounds as substrate. Among the substrates used, the greatest substrate specificity was observed with catechol. Durum wheat PPO was sensitive to inhibitors such as ascorbic acid, cysteine, oxalic acid and citric acid. Ascorbic acid was the most effective inhibitor.     

The contribution of ear photosynthesis to grain filling in bread wheat (Triticum aestivum L.)

The contribution of ear photosynthesis to grain filling in wheat (Triticum aestivum L.) is not well known. The main objective of this work was to evaluate this contribution through three different experimental approaches: (1) ear photosynthesis was reduced by removing awns or shading the ears (in combination with a defoliation treatment), (2) grain weight per ear was compared in an ‘all shaded’ crop versus plants where only the vegetative parts were shaded (‘ear emerging’), and (3) ear photosynthesis was reduced with DCMU (3-(3,4-dichlorophenyl)-1,1-dimethylurea), a specific inhibitor of photosystem II.     

青海和西藏小麦品种主要春化基因的组成分析

为了明确青海和西藏小麦春化基因的分布特点,利用STS标记对96个青海和西藏小麦品种主要春化基因VRN-A1、VRN-B1、VRN-D1和VRN-B3位点的等位变异组成进行了检测和分析。在96个小麦品种中,VRN-A1位点存在Vrn-A1 a(30.2%)、Vrn-A1 b(6.3%)和vrn-A1(63.5%)3种等位变异,VRN-B1位点存在Vrn-B1 a(29.2%)、Vrn-B1 b(12.5%)和vrn-B1(58.3%)3种等位变异,VRN-D1位点存在Vrn-D1 a(55.2%)、Vrn-D1 b(1.0%)和vrn-D1(43.8%)3种等位变异,VRN-B3位点存在Vrn-B3 b(1.0%)和vrn-B3(99.0%)两种等位变异。4个位点显性春化基因等位变异的分布频率不同,依次为Vrn-D1(56.3%)Vrn-B1(41.7%)Vrn-A1(36.5%)Vrn-B3(1.0%)。在14个冬小麦品种中,4个位点均为隐性春化基因等位变异;在82个春小麦品种中,4个位点至少有1个携带显性等位变异,其中VRN-D1位点显性等位变异占主导地位,并常与其他位点显性等位变异伴随出现。两个地区春小麦品种间,4个位点显性等位变异的分布频率存在较大的差异,青海春小麦品种依次为Vrn-B1(64.8%)Vrn-A1(51.9%)=Vrn-D1(51.9%)Vrn-B3(1.9%),西藏春小麦品种为Vrn-D1(92.9%)Vrn-A1(25.0%)Vrn-B1(17.9%)Vrn-B3(0)。在82个春小麦品种中,4个春化基因位点存在8种等位变异组合类型,vrn-A1/vrn-B1/Vrn-D1/vrn-B3(30.2%)Vrn-A1/Vrn-B1/vrn-D1/vrn-B3(17.6%)vrn-A1/Vrn-B1/Vrn-D1/vrn-B3(11.5%)Vrn-A1/vrn-B1/Vrn-D1/vrn-B3(7.3%)vrnA1/Vrn-B1/vrn-D1/vrn-B3(6.3%)=Vrn-A1/Vrn-B1/Vrn-D1/vrn-B3(6.3%)Vrn-A1/vrn-B1/vrn-D1/vrn-B3(5.2%)vrn-A1/vrn-B1/Vrn-D1/Vrn-B3(1.0%)。这8种等位变异组合在春小麦中的分布频率因品种推广地区不同而不同。在青海春小麦品种中,存在8种等位变异组合类型,以Vrn-A1/Vrn-B1/vrn-D1/vrn-B3(29.5%)和vrn-A1/vrn-B1/Vrn-D1/vrn-B3(22.2%)为主,vrn-A1/Vrn-B1/Vrn-D1/vrn-B3(14.8%)与Vrn-A1/Vrn-B1/Vrn-D1/vrn-B3(11.1%)次之;在西藏春小麦品种中,仅存在5种等位变异组合类型,以vrn-A1/vrn-B1/Vrn-D1/vrn-B3(60.7%)为主,Vrn-A1/vrn-B1/Vrn-D1/vrn-B3(21.4%)和vrn-A1/Vrn-B1/Vrn-D1/vrn-B3(10.7%)次之,vrn-A1/Vrn-B1/vrn-D1/vrn-B3(3.6%)和Vrn-A1/Vrn-B1/vrn-D1/vrn-B3(3.6%)较低。这些信息可为青海和西藏小麦品种选育和推广提供依据。