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Changes in specialized metabolites were analyzed in wheat leaves inoculated with Bipolaris sorokiniana, the causal agent of spot blotch of Poaceae species. HPLC analysis detected the accumulation of six compounds in B. sorokiniana-infected leaves. Of these, we purified two compounds by silica gel and ODS column chromatography and preparative HPLC, and identified them as cinnamic acid amides, N-cinnamoyl-9-hydroxy-8-oxotryptamine and N-cinnamoyl-8-oxotryptamine, by spectroscopic analyses. The remaining four compounds were predicted to be p-coumaric acid amides of hydroxyputrescine, hydroxyagmatine, hydroxydehydroagmatine, and agmatine by mass spectrometry. The accumulation of two cinnamic acid amides was also induced by Fusarium graminearum infection, and by treatment with CuCl2, jasmonic acid, and isopentenyladenine. Antifungal activity of these amides was shown by inhibition of conidial germination and germ tube elongation of F. graminearum and Alternaria brassicicola, indicating that they act as phytoalexins. The accumulation of these amides also detected in barley leaves treated with CuCl2. We examined the accumulation of 25 phenylamides in B. sorokiniana-infected wheat leaves using LC-MS/MS. Hydroxycinnamic acid amides of tryptamine, serotonin, putrescine, and agmatine, were induced after infection with B. sorokiniana. Thus, the induced accumulation of two groups of phenylamides, cinnamic acid amides with indole amines, and p-coumaric acid amides with putrescine and agmatine related amines, represents a major metabolic response of wheat to pathogen infection.

(1,3;1,4)-β-D-glucans (mixed-linkage glucans) are found in tissues of members of the Poaceae (grasses), and are particularly high in barley (Hordeum vulgare) grains. The present study describes the isolation of three independent (1,3;1,4)-β-D-glucanless (betaglucanless; bgl) mutants of barley which completely lack (1,3;1,4)-β-D-glucan in all the tissues tested. The bgl phenotype cosegregates with the cellulose synthase like HvCslF6 gene on chromosome arm 7HL. Each of the bgl mutants has a single nucleotide substitution in the coding region of the HvCslF6 gene resulting in a change of a highly conserved amino acid residue of the HvCslF6 protein. Microsomal membranes isolated from developing endosperm of the bgl mutants lack detectable (1,3;1,4)-β-D-glucan synthase activity indicating that the HvCslF6 protein is inactive. This was confirmed by transient expression of the HvCslF6 cDNAs in Nicotiana benthamiana leaves. The wild-type HvCslF6 gene directed the synthesis of high levels of (1,3;1,4)-β-D-glucans, whereas the mutant HvCslF6 proteins completely lack the ability to synthesize (1,3;1,4)-β-D-glucans. The fine structure of the (1,3;1,4)-β-D-glucan produced in the tobacco leaf was also very different from that found in cereals having an extremely low DP3/DP4 ratio. These results demonstrate that, among the seven CslF and one CslH genes present in the barley genome, HvCslF6 has a unique role and is the key determinant controlling the biosynthesis of (1,3;1,4)-β-D-glucans. Natural allelic variation in the HvCslF6 gene was found predominantly within introns among 29 barley accessions studied. Genetic manipulation of the HvCslF6 gene could enable control of (1,3;1,4)-β-D-glucans in accordance with the purposes of use.

A QTL analysis was performed to determine the inheritance of flowering time in barley, using a set of recombinant inbred lines developed from a winter-type × spring-type cross. Two photoperiod responsive loci, Ppd-H1 and Ppd-H2, were detected on chromosome arms 2HS and 1HL respectively. Segregation for eam8 (mapping to the terminus of chromosome arm 1HL) and Eam5 (close to Sgh2 on chromosome arm 5HL) was also observed. These latter two genes functioned under 12 h to 24 h photoperiods. In addition, eps2S and eps7S, known to lie on chromosome arms 2HS and 7HS respectively, were detected. A new QTL for flowering time, qDHE.ak-1HS, was mapped 23 cM from the terminus of chromosome 1HS, and appears to be expressed under extremely short day lengths.

Shiratae Nijo is a new two-rowed food barley cultivar bred at the National Agricultural Research Center for Kyushu Okinawa Region (KONARC) in collaboration with the National Institute of Crop Science (NICS) and released in 2009. Shiratae Nijo is a proanthocyanidin-free cultivar bred by backcross breeding using a leading cultivar Nishinohoshi as a recurrent parent and a foreign proanthocyanidin-free mutant ant28-494 as a non-recurrent parent. The agronomic characteristics of Shiratae Nijo are almost the same as those of Nishinohoshi which has high yield performance and high resistance to barley yellow mosaic virus (BaYMV) type I and powdery mildew. Like Nishinohoshi, Shiratae Nijo shows high resistance to Fusarium head blight (FHB) at the anthesis stage but is susceptible 10 days after anthesis. Shiratae Nijo exhibits significantly weaker seed dormancy than Nishinohoshi. Shiratae Nijo has excellent pearling quality, and the pearled grains show no discoloration and high whiteness after cooking because of the lack of catechin and proanthocyanidin in the grains.

黒ボク土など穀粒が高タンパク質化しやすい栽培環境における精麦品質の向上に有用な形質を明らかにするため，胚乳が粉状質を呈するモチ性および破砕デンプン粒変異（fractured starch granule）に関する準同質遺伝子系統を用いて，灰色低地土水田と黒ボク土畑における硝子率と精麦品質の差異を解析した．その結果，これらの準同質遺伝子系統ではタンパク質含量にかかわらず，ほぼ完全な粉状質であり，黒ボク土畑においても硝子質粒がほとんど発生しないことが明らかになった．また，モチ性および破砕デンプン粒変異の準同質遺伝子系統は，栽培土壌にかかわらず原系統よりも高白度であった．搗精時の砕粒発生は原系統と同程度かむしろ少なく，胚乳が粉状質であっても砕けやすい脆い粒質ではないことが明らかとなった．以上のことから，モチ性や破砕デンプン粒変異は，高タンパク質化の影響を受けない粉状質高白度品種の育成において有用な形質であると考えられた．また，これらの準同質遺伝子系統ではデンプン含量が減少したが，胚乳および糊粉層の細胞壁多糖であるβ–グルカンの含量が増加することが認められた．β–グルカンは，人体にとって種々の機能性を有する多糖であるため，モチ性や破砕デンプン粒変異は食用オオムギの付加価値の向上においても有用な形質であると考えられた．とくに，破砕デンプン粒変異はウルチ性の完全粉状質品種の育成に有用な形質であると考えられた．

(1-3, 1-4)-β-D-glucan contained in barley (Hordeum vulgare L.) grains is a main component of endosperm cell walls and constitutes the cell wall matrix with arabinoxylan. A monofactorial recessive mutant gene for (1-3, 1-4)-β-D-glucanless grain was found and the new gene was designated as bgl (= (1-3, 1-4)-beta-D-glucanless grain). A linkage was found between the bgl gene and the naked caryopsis (nud) gene, and it was mapped to the centromeric region of chromosome 7H. Phenotypes in bgl cosegregated with the polymorphisms in HvCslF6, a member of cellulose synthase-like HvCslF gene family, indicating that bgl was caused by a mutation in the HvCslF6 locus. In order to clarify the characteristics of (1-3, 1-4)-β-D-glucanless barley grains, a near-isogenic line (NIL) was developed by backcrossing the Japanese two-rowed cultivar ‘Nishinohoshi’ as a recurrent parent. The NIL with bgl completely lacked (1-3, 1-4)-β-D-glucan in both the endosperm and aleurone layer cell walls. Microscopic analysis revealed that the NIL had thin endosperm cell walls. It also showed a softer grain texture and many more broken grains during the pearling process than the recurrent parent. The soft and friable grain texture of the NIL was probably caused by the thin endosperm cell walls.

In the western part of Japan, two wheat cultivars, Nishinokaori and Minaminokaori, are currently cultivated for breadmaking. Breadmaking wheat requires a higher protein content compared to the Japanese noodle wheat (the major type of wheat in Japan). This high protein level in the grain is obtained by top-dressing with nitrogen (N) near anthesis. Because such N applications may increase levels of Fusarium head blight (FHB) and consequent mycotoxin [deoxynivalenol (DON) and nivalenol (NIV)] accumulation in the grain, the effect of N application (0, 4, and 8 g/m2) at anthesis on FHB and mycotoxin accumulation in Nishinokaori and Minaminokaori was tested in the greenhouse in 2004 and 2005 and in two fields in 2006. In the greenhouse, plants were spray inoculated at 3, 10, and 20 days after N treatment. In field experiments, colonized maize kernels, which generate ascospores during the testing season, served as inoculum. In all experiments for both cultivars, N application at anthesis significantly increased grain protein as expected, but had no significant effect on FHB and DON and NIV levels in grain. These results suggest that, at least in these cultivars, N can be applied close to anthesis without increasing the risk of FHB and mycotoxin (DON and NIV) accumulation.

Flavonoids accumulated in proanthocyanidin-free near-isogenic lines iso ant 13, iso ant 17, and iso ant 22 of Nishinohoshi, developed by backcross breeding using a leading cultivar, Nishinohoshi, as a recurrent parent and a proanthocyanidin-free mutant as a nonrecurrent parent in Japan, were examined. A new flavanone, (2RS)-dihydrotricin 7-O-β-D-glucopyranoside (1), known flavanones (2RS)-dihydrotricin (2) and (2RS)-homoeriodictyol (3), and known flavones chrysoeriol 7-O-[α-L-rhamnopyranosyl-(1→6)-β-D-glucopyranoside] (4), chrysoeriol 7-O-β-D-glucopyranoside (5), tricin (6), and chrysoeriol (7) were isolated from iso ant 17 of Nishinohoshi. The structures and stereochemistries of the isolated flavonoids (1-7) were elucidated on the basis of spectroscopic analyses. The concentrations of the isolated flavonoids (1-7) in iso ant 13, iso ant 17, and iso ant 22 of Nishinohoshi were similar to each other, whereas the flavonoids 1-5 and 7 were not detected in Nishinohoshi, an old Japanese cultivar, Amaginijo, and North American cultivar Harrington. The concentration of tricin (6) in Nishinohoshi was a half those in iso ant 13, iso ant 17, and iso ant 22 of Nishinohoshi. Except for iso ant 13, iso ant 17, and iso ant 22 of Nishinohoshi, the concentration of tricin (6) was highest in Nishinohoshi, followed by Amaginijo and Harrington. Thus, tricin (6), its precursor dihydrotricin (2), and its glucopyranoside, dihydrotricin 7-O-β-D-glucopyranoside (1), as well as chrysoeriol (7) and homoeriodictyol (3) were accumulated in iso ant 13, iso ant 17, and iso ant 22 of Nishinohoshi probably by blocking at the step of flavanone 3-hydroxylase in the procyanidin biogenetic pathway, resulting in enhancement of the alternative biogenetic pathway.

Five barley (Hordeum vulgare) PEBP (for phosphatidylethanolamine-binding protein) genes were analyzed to clarify their functional roles in flowering using transgenic, expression, and quantitative trait locus analyses. Introduction of HvTFL1 and HvMFT1 into rice (Oryza sativa) plants did not result in any changes in flowering, suggesting that these two genes have functions distinct from flowering. Overexpression of HvFT1, HvFT2, and HvFT3 in rice resulted in early heading, indicating that these FT-like genes can act as promoters of the floral transition. HvFT1 transgenic plants showed the most robust flowering initiation. In barley, HvFT1 was expressed at the time of shoot meristem phase transition. These results suggest that HvFT1 is the key gene responsible for flowering in the barley FT-like gene family. HvFT2 transgenic plants also showed robust flowering initiation, but HvFT2 was expressed only under short-day (SD) conditions during the phase transition, suggesting that its role is limited to specific photoperiodic conditions in barley. Flowering activity in HvFT3 transgenic rice was not as strong and was modulated by the photoperiod. These results suggest that HvFT3 functions in flowering promotion but that its effect is indirect. HvFT3 expression was observed in Morex, a barley cultivar carrying a dominant allele of Ppd-H2, a major quantitative trait locus for flowering under SD conditions, although no expression was detected in Steptoe, a cultivar carrying ppd-H2. HvFT3 was expressed in Morex under both long-day and SD conditions, although its expression was increased under SD conditions. HvFT3 was mapped to chromosome 1HL, the same chromosome that carries Ppd-H2. Genomic sequence analyses revealed that Morex possesses an intact HvFT3 gene, whereas most of this gene has been lost in Steptoe. These data strongly suggest that HvFT3 may be identical to Ppd-H2.