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Heading/flowering at the appropriate time in each cultivation region can greatly contribute to stabilizing and maximizing yield. Genotype combinations of Vrn-1 and Ppd-1, the major genes determining heading time, can explain the variation in heading time. However, the genes that can explain the remaining variations in heading time are still unknown. In this study, we aimed to identify the genes conferring early heading using doubled haploid lines derived from Japanese wheat varieties. Quantitative trait loci (QTL) analysis revealed a significant QTL on the long arm of chromosome 1B in multiple growing seasons. Genome sequencing using Illumina short reads and Pacbio HiFi reads revealed a large deletion of a ~500 kb region containing TaELF-B3, an orthologue of Arabidopsis clock gene EARLY FLOWERING 3 (ELF3). Plants with the deleted allele of TaELF-B3 (ΔTaELF-B3allele) headed earlier only under short-day vernalization conditions. Higher expression levels of clock- and clock-output genes, such as Ppd-1 and TaGI, were observed in plants with the ΔTaELF-B3 allele. These results suggest that the deletion of TaELF-B3 causes early heading. Of the TaELF-3 homoeoalleles conferring early heading, the ΔTaELF-B3 allele showed the greatest effect on the early heading phenotype in Japan. The higher allele frequency of the ΔTaELF-B3 allele in western Japan suggests that the ΔTaELF-B3 allele was preferred during recent breeding to adapt to the environment. TaELF-3 homoeolog will help to expand the cultivated area by fine-tuning the optimal timing of heading in each environment.

Cadmium (Cd) is as an extremely toxic metal that can contaminate agricultural soils. To reduce the risk of Cd intake in food cereals, the development of cultivars with low grain Cd concentration (GCC) is an effective countermeasure. We analyzed quantitative trait loci (QTLs) for GCC in a doubled haploid (DH) common wheat (Triticum aestivum L.) population derived from ‘Chugoku 165’ (low GCC) × ‘Chukei 10-22’ (high GCC). We found novel loci for low GCC on the short arm of chromosome 4B and on the long arm of chromosome 6B. These QTLs accounted for 9.4%–25.4% (4B) and 9.0%–17.8% (6B) of the phenotypic variance in the DH population. An association analysis with 43 cultivars identified 3 loci at these QTLs: QCdc.4B-kita, QCdc.6B-kita1, and QCdc.6B-kita2. In contrast to durum wheat and barley, no QTL was detected on the chromosomes of homeologous group 5 for heavy metal P1B-type ATPase 3. These results will contribute to marker-assisted selection for low GCC in breeding of common wheat.

Preharvest sprouting (PHS) of durum wheat (Triticum turgidum ssp. durum (Desf.) Husn.) is an important problem in Japan, where the rainy season overlaps with the harvest season. Since there are few PHS-resistant genetic resources in durum wheat, we introduced an R-gene for red seeds, the MFT gene, and the QPhs-5AL QTL, all of which are associated with PHS resistance, into durum wheat from a PHS-resistant bread wheat (T. aestivum L.) cultivar, ‘Zenkoujikomugi’ (Zen), by backcross breeding. Developed near isogenic lines (NILs) with red seeds had a lower percentage germination (PG) and germination index (GI) than the recurrent parent, and seed color had the greatest effect. A NIL combining all three sequences had the lowest GI and PG, with a similar GI to that of ‘Shiroganekomugi’ bread wheat. Among NILs with white seeds, a NIL combining MFT and QPhs-5AL had the lowest GI and PG. As the combination of all three sequences from Zen conferred PHS resistance on durum wheat, PHS-resistant genetic resources in bread wheat can be used in breeding durum wheat.

Chloride (Cl) in saline soil increases the cadmium (Cd) concentration in crops. Here, we conducted a field experiment to investigate changes in Cd concentrations in wheat grain after the application of the Cl-containing fertilizer ammonium chloride (NH4Cl), with the aim of reducing its potential health risk. Effects of the application stage of NH4Cl fertilizer and leaching treatment (i.e., heavy rainfall) were also investigated in field and pot experiments. Both field and pot experiments showed that the Cd concentration of wheat grain was higher with NH4Cl fertilizer than with ammonium sulfate or urea fertilizers. Grain Cd concentration in wheat fertilized with NH4Cl at the tillering–jointing and flowering stages in the field experiment was 0.223 mg kg–1, which was about 1.5 times higher than that fertilized with urea. This finding is important because, in Japan, compound fertilizers containing NH4Cl are commonly used in fields for wheat cultivation. NH4Cl fertilizer application at the tillering–j...

Grain hardness, amylose content, and glutenin subunit composition are critical determinants for end-use properties of wheat. To improve the end-use properties of domestic wheats, we studied these traits between the Australian and North American wheat classes exported to Japan in 2009 and 2011 by analyzing the corresponding alleles. Most hard classes had Pina-D1b or Pinb-D1b. A partial waxy allele (Wx-B1b) was found in all Australian Standard White (ASW) seeds in 2009 and two-thirds of ASW seeds in 2011. All or most American hard wheat seeds had Glu-D1d. Most U.S. Western White (WW) seeds had a null allele (Glu-A1c) or alleles that lacked one of the two Glu-B1 subunits. Most hard red winter (HRW) seeds had Glu-B3b or Glu-B3g. Quality characteristics of these classes seemed to be consistent with these results. In addition, we also found new Glu-1 and Glu-3 alleles in HRW and WW. These results suggested that although there are variations in its allelic composition from year to year, each class has u...

To study the influence of genes from Thinopyrum intermedium on traits affecting the bread-making quality of wheat, two derivatives from a putative disomic addition line in cultivar ‘Vilmorin 27’ were used in cytological, biochemical and molecular characterization. Cytological analysis suggested that one of the derivatives (Line-1) had a terminal deletion involving the long arm of chromosome 1D (2n = 42, Del-1DL”), and the other (Line-2) was a conventional addition line, but also carried the same deletion on chromosome 1D (2n = 44, Thi”+Del-1DL”). Amplification and sequencing of high-molecular-weight glutenin subunit (HMW-GS) genes coded by the Th. intermedium chromosome in Line-2 indicated the presence of one x-type with an extra cysteine and four (rather than one) unique y-type genes. Rheological studies of Line-1 showed significantly lower dough strength compared to ‘Vilmorin 27’, confirming the recognized role of Glu-1D coded HMW-GSs. Line-2 showed significantly higher dough strength compared to the background cultivar, indicating a significant potential of Th. intermedium for improvement of bread-making quality in wheat.

Phytoextraction using indica rice plants (Indian Rice Oryza ssp.) is a promising technique for remediating cadmium (Cd)-polluted paddy fields. Because this technique has only been established for paddy fields, we decided to examine phytoextraction in upland fields that have been converted from paddy fields. Although “CHOKOUKOKU” shows a shattering habit and lodging, its Cd uptake was significantly higher than that of other indica rice varieties. On the other hand, “IR8”, which was able to accumulate only moderate levels of Cd in its shoots, has a lodging tolerance, making it an optimal variety for southwest Japan, which experiences several typhoons each year. Therefore, both “CHOKOUKOKU” and “IR8” were useful in estimating practical phytoextraction in upland fields. A practical phytoextraction examination showed that the total Cd uptake of “CHOKOUKOKU” and “IR8” was 822 and 545 g ha−1, respectively, after a 4-year period. After phytoextraction by planting high Cd-accumulating rice plants, the Cd c...

Jasmonates, including jasmonic acid and its derivatives such as methyl jasmonate (MeJA), are plant growth substances that control various responses. Jasmonates regulate leaf trichome density in dicotyledonous plants, but their effects on the trichome density of monocotyledonous plants, such as those in the Poaceae, remain unclear. In the present study we examined the effects of exogenous MeJA on the trichome density of Rhodes grass, which has three kinds of trichomes: macrohairs, salt glands, and prickles. Exogenous MeJA significantly increased the densities of macrohairs and salt glands on the adaxial and abaxial leaf surfaces and those of prickles on the adaxial leaf surface. Because exogenous MeJA significantly reduced the leaf area, we calculated the number of trichomes per 1000 epidermal cells to eliminate the effects of reduced leaf area. Exogenous MeJA significantly increased the number of macrohairs per 1000 epidermal cells on both adaxial and abaxial leaf surfaces, but it significantly decreased the number of salt glands per 1000 epidermal cells on both surfaces. Exogenous MeJA had no significant effects on the number of prickles per 1000 epidermal cells on either of the leaf surfaces. These results indicate that exogenous MeJA alters the trichome density by affecting leaf area and trichome initiation, and the effects of exogenous MeJA on trichome initiation differ among the various trichome types.

After screening of 177 disomic addition lines (DALs) of wheat (Triticum aestivum) containing a pair of chromosomes from different alien species, we found that the chromosome 1E addition line of Agropyron elongatum, that is known to be a potential genetic resource for drought and salinity tolerance, showed potential for improvement of bread-making quality of wheat. This was indicated by increased SDS sedimentation, specific sedimentation, mixograph peak time and SE-HPLC analysis of polymeric proteins. This addition line spontaneously gave rise to a substitution line for chromosome 1D in subsequent generations that showed weak dough strength. Analysis of the x-type HMW-glutenin subunit sequence of Ag. elongatum from DAL1E indicated that it closely resembled the x-type sequence of the A and B genomes of wheat, and the y-type was intermediate between x- and y-type HMW-glutenin subunit genes. From these observations, it was inferred that 1E-encoded seed storage proteins have considerable potential for improvement of wheat end-product quality if transferred to specific chromosomes such as 1A of Chinese Spring (CS) wheat, which has a negative overall effect on bread-making quality.

Using various rice flours prepared by different milling methods, the relationship between rice flour properties and specific loaf volumes of one-loaf bread made from rice flour with wheat vital gluten were studied. Damaged starch content of rice flour varied from 1.0% to 22.1%. A significant negative correlation was verified between damaged starch content and specific loaf volume of one-loaf bread. Rice flour with low damaged starch content mainly consisted of compound starch granules, aggregated polyhedral single starch granules and smooth surface cells surrounded by the cell wall. The structures of starch granules and cells were maintained in rice flour with low damaged starch content. Rice flour with high damaged starch content consisted of only fine irregular particles without the apparent rice starch structure, or contained fractured large and small cells with rough surface. Although there was not significant correlation between specific loaf volume and median particle size, rice flours which were successful at making bread with high specific loaf volume commonly showed a peak centered around 60 µm with a smaller amount of larger size particles in the particle distribution profile. Thus, the flour particle size distribution appears to affect the specific loaf volume of one-loaf bread. Both the damaged starch content and the profile of particle size distribution were important for high specific loaf volume of one-loaf bread. Less damage to starch and cell structures while lowering the particle size during milling process is critical in obtaining better flour for rice one-loaf bread making.

This study involved screening of wild species of wheat in search of functionally useful seed storage proteins for improvement of breadmaking quality of wheat (Triticum aestivum). After screening of 177 disomic addition lines (DALs) of wheat belonging to different wild species, Aegilops searsii DALs were selected and studied in detail. These DALs of Ae. searsii were from chromosome 1Ss to 7Ss in the background of cultivated wheat cv. Chinese Spring (CS). By analyzing these addition lines, genetic loci of actively expressed genes for the high molecular weight glutenin subunits (HMW-GS) and gliadin were found on the chromosome 1Ss for the first time and have been designated as Glu-Ss1 and Gli-Ss1, respectively. Disomic addition line of chromosome 1Ss (DAL1Ss) showed improved dough strength in different generations compared with CS. SDS sedimentation value and specific sedimentation of DAL1Ss were higher than CS. Mixograph peak height and band width were higher, with no difference in mixing peak time...

A “two-line system” using photoperiod-sensitive cytoplasmic male sterility (PCMS) caused by Aegilops crassa cytoplasm under long-day photoperiods (≧15 h) has been proposed as a means of producing hybrid varieties in common wheat (Triticum aestivum). The PCMS line is maintained by self-pollination under short-day conditions, and hybrid seeds can be produced through outcrossing of the PCMS line with a pollinator line under long-day conditions. Our previous studies revealed that PCMS lines showing complete male sterility under long-day conditions are necessary for practical hybrid wheat breeding, especially to obtain high hybrid purity in F1 seeds. Furthermore, practical PCMS lines should have high seed fertility under short-day conditions, which is associated with female fertility. Wheat cv. Norin 26 with Ae. crassa cytoplasm exhibits high seed fertility under short-day conditions, and cv. Fujimikomugi with Ae. crassa cytoplasm shows high male sterility under long-day conditions. Here we developed practical PCMS lines derived from the F1 generation of Norin 26 and Fujimikomugi (with Ae. crassa cytoplasm) that were then backcrossed to elite wheat lines.

Allelic variation of high molecular weight glutenin subunits (HMW-GS) encoded by Glu-A1, Glu-B1 and Glu-D1 in wheat landraces originating from the Xinjiang Uygur Autonomous District (Xinjiang District) of China was investigated by SDS-PAGE and the relationship between the variation and the winter habit was examined. The most frequent allele at the Glu-A1 locus was Glu-A1c (null allele: 75.5%), followed by Glu-A1b encoding subunit 2* (22.3%). These two alleles were distinctive at Glu-A1 in the landraces of both spring and winter cultivars. No difference in allele frequency was found between the spring and the winter wheat cultivars. At the Glu-B1 locus, the most frequent allele was Glu-B1b (90.8%) encoding subunits 7 + 8, the allele frequency of which was almost the same between the spring and the winter wheat cultivars. Glu-D1a (72.0%) encoding subunits 2 + 12 was a major allele at the Glu-D1 locus among the landraces. Although no cultivar with Glu-d1f encoding subunits 2.2 + 12 was found, the novel subunit pair 2.6 + 12 (24.5%) was observed only in the winter wheat cultivars. Subunit 2.6 was slightly less mobile than subunit 2.2. The major Glu-D1 allele was Glu-D1a (94.5%) among the spring wheat cultivars and this allele characterized the spring wheat cultivars in the Xinjiang District. Both the subunit pair 2.6 + 12 encoded by the Glu-D1bp(t) allele (58.5%) and the subunit pair 2 + 12 encoded by the Glu-D1a allele (40.7%) predominated among the winter wheat cultivars. The large difference in allelic variation at the Glu-D1 locus suggested that the origin of the spring wheat cultivars was different from that of the winter wheat cultivars. The absence of landrace with Glu-D1f suggested that Xinjiang wheat was not related to Japanese wheat.

We studied the effects of hardness allele Pinb-D1b on the flour quality characters for Japanese white salty noodles. With 110 doubled haploid lines derived from ‘Chugoku 140’ (hard, Pinb-D1b )בChikugoizumi’ (soft, Pinb-D1a ), Pinb-D1b hard lines were found to have larger L* and a* of 60% flour colour, and smaller RVA breakdown than those of Pinb-D1a soft lines in addition to the higher flour yield, and larger average flour particle size. The Pinb-D1b allele is thus regarded to effectively improve milling efficiency and flour brightness. The inferior flour viscosity of Pinb-D1b lines is compensated for by introducing waxy allele(s) to increase viscosity by lowering amylose content.

Grain hardness is an important factor affecting end-use quality in wheat. Mutations of the puroindoline genes, which are located on chromosome 5DS, control a majority of grain texture variations. Hordoindoline genes, which are the puroindoline gene homologs in barley, are located on chromosome 5HS and are also responsible for grain texture variation. In this study, we used three types of wheat–barley species (Hordeum vulgare, H. vulgare ssp. spontaneum, and H. chilense) chromosome addition lines and studied the effect of chromosome 5H of these species on wheat grain characteristics. The 5H chromosome addition lines showed significantly lower grain hardness and higher grain weight than the corresponding wheat parents. The effect of enhancing grain softness was largest in the wheat–H. chilense line regardless of having an increase in grain weight similar to those in the wheat–H. vulgare and wheat–H. spontaneum lines. Our results indicated that chromosome 5H of the Hordeum species plays a role in enhancing grain softness and increasing grain weight in the wheat genetic background, and the extent of effect on grain hardness depends on the type of Hordeum species. Protein analysis of hordoindolines indicated that profiles of 2D-electrophoresis of hordoindolines were different among Hordeum species and hordoindolines in the addition lines appeared to be most abundant in wheat–H. chilense line. The differences in enhancing grain softness among the Hordeum species might be attributed to the quantity of hordoindolines expressed in the 5H chromosome addition lines. These results suggested that the barley hordoindolines located on chromosome 5HS play a role in reducing grain hardness in the wheat genetic background.

The production of angiotensin I converting enzyme (ACE) inhibitory peptides by autolysis reactions from wheat milling byproducts was investigated. Milled whole grain, bran, shorts, and red dog acquired ACE inhibitory activity though water soaking treatment. Among the milled fractions, the preparation of shorts exhibited the strongest inhibitory activity (IC50 = 0.08 mg protein/mL) followed by that of bran, red dog, and whole grain in decreasing order. The production of ACE inhibitory peptides was almost completely inhibited by pepstatin A, indicating the contribution of aspartic proteases. The optimal pH for acquiring ACE inhibitory activity of the byproduct fraction (mixtures of bran and shorts) was 3.2. The level of inhibitory activity rose with increasing temperature up to 40 degrees C. The inhibitory activity reached a maximal level after a 12 h reaction time and maintained the same level up to 24 h at 40 degrees C, pH 3.2. From the hydrolysate of the byproduct fraction, six peptides were isolated by several steps of chromatography, and their amino acid sequences were Leu-Gln-Pro, Ile-Gln-Pro, Leu-Arg-Pro, Val-Tyr, Ile-Tyr, and Thr-Phe. Thus, wheat milling byproducts have the possibility to become an effective source for ACE inhibitory peptides.

The methylotrophic yeasts are commonly used as hosts for heterologous gene expression. In this study, we describe a novel gene, TRM1 , in Candida boidinii , responsible for the transcriptional activation of several methanol-inducible promoters. The encoded protein, Trm1p, is a Zn(II) 2 Cys 6 -type zinc cluster protein. Deletion of TRM1 completely inhibits growth on methanol but causes no growth defect on glucose or other nonfermentative carbon sources, glycerol, ethanol, or oleate. Trm1p is responsible for transcriptional activation of five methanol-inducible promoters tested, but not for peroxisome assembly or peroxisomal protein transport. Expression of the TRM1 gene was constitutive, and Trm1p localizes to the nuclei regardless of the carbon source. Two cis -acting methanol response elements (MREs), MRE1 and MRE2 are present in the promoter of the dihydroxyacetone synthase gene. Trm1p is shown to be required for MRE1-dependent methanol-inducible gene expression. Chromatin immunoprecipitation assays reveal that Trm1p binds to five methanol-inducible promoters upon methanol induction but does not bind in glucose-grown cells. Thus, the TRM1 gene encodes a master transcriptional regulator responsible for methanol-specific gene activation in the methylotrophic yeasts.