Brown-hilum soybeans often have cold-weather tolerance and growth vigor superior to that of yellow hilum soybeans. However, Japanese consumers do not prefer brown-hilum soybeans. In this paper, we investigated the correlation of cold-weather tolerance with pubescence color and flowering time using near-isogenic lines (NILs). The gene for the former is one of the genes, which controls hilum color and that for the latter is related to growth vigor. First, the NILs for pubescence color (tawny: T and gray: t) of yellow-hilum soybeans were grown under various temperature conditions, and their growth and seed yield were examined. There was no significant difference in the seed yield between tawny and gray lines in a warm region for eastern Hokkaido, but the seed yield in tawny lines was significantly higher than that in gray lines in cooler regions. And dry matter production at the flowering stage was also higher in tawny lines than in gray lines in cooler regions. Next, the effects of cooling treatment on the pod number and seed yield of the NILs for a combination of pubescence color (T/t) and flowering time (E1/e1), te, tE, Te and TE, were examined. The cooling treatment [four weeks, at 18°C/13°C (day/night)] from the beginning of flowering slightly reduced the pod number and seed yield in the tawny lines (Te, TE) and in the late-flowering gray line (tE), but greatly reduced them in the early flowering gray line (te). Then, gray pubescence lines were grouped into early, mid and late flowering groups from two crosses, and they were exposed to low temperature mentioned above. The rates of reduction in the pod number and seed yield by the cooling treatment were highest (cold-weather tolerance was lowest) in the earlier flowering group and lowest (cold-weather tolerance was highest) in the later flowering group. These results show that selecting yellow-hilum soybeans with tawny pubescence and with later flowering time are both useful for developing yellow-hilum soybeans with cold-weather tolerance.

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We conducted diallel analyses of root shape and size in Japanese radish to determine the interaction between genetic effects and soil type in the selection for these traits in a breeding program. A set of diallel crosses of six inbred lines was grown in an andosol, a clayey soil and a sandy soil. The major shape characteristics accounted for by the principal components of elliptic Fourier descriptors were the ratio of length to width, the bluntness of the distal part, the curvature of the central part, and the curvature of the proximal and distal parts. Analysis of variance of diallel tables showed that the genetic effects were highly significant for all the characteristics. In contrast, the influence of the soil type was markedly different among the characteristics. Soil type effect was significant for size characteristics and the bluntness of the distal part. In the bluntness of the distal part, the interaction between genetic effects and soil type was significant, and the heritability and the degree of dominance varied between the soil types. The results indicated that the differences in the soil types did not influence the selection for the root size or shape in most characteristics. However, the interaction between genetic effects and soil type should be taken into account in the selection for the bluntness of the distal part.

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A Chinese landrace of barley, Mokusekko 3, is completely resistant to all the present strains of Barley yellow mosaic virus (BaYMV) and Barley mild mosaic virus (BaMMV), and has at least two resistance genes, rym1 and rym5. However, rym1 has not been used for BaYMV-resistance breeding of barley in Japan so far. In our previous study, rym1 was completely resistant to BaYMV-I, -II, BaMMV-Ka1 and -Na1, and was acceptably resistant to BaYMV-III. Complete resistance of Mokusekko 3 was estimated to be due to the interaction between rym1 and rym5. To estimate pyramiding effects of these genes using DNA markers, we investigated the resistance reaction of lines with or without these genes to BaYMV-I and -III. All the pyramiding lines showed complete resistance to BaYMV-I and -III. This study anticipated the resistance to BaYMV-III of Mokusekko 3 by the multiple effect of the rym1 and rym5 genes.

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The present study was carried out to identify QTLs controlling heading time of barley. A total of 89 recombinant inbred lines (RILs) generated from a cross between the Oriental-type winter cultivar ‘Azumamugi’ and the Occidental-type spring cultivar ‘Kanto Nakate Gold’ were examined. Two field experiments consisting of one spring sowing and one autumn sowing and a greenhouse experiment under 24 h illumination were conducted. Two genes for the spring habit and five QTLs controlling heading time were identified. The two genes for the spring habit, sgh1 and Sgh2 were located on chromosomes 4HL and 5HL, respectively; four QTLs were mapped close to the heading time genes identified in the previous studies on chromosomes 1HL, 2HS, 2HL and 7HS; and one was a new QTL detected on chromosome 7H.

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The sd1 (dee-geo-woo-gen dwarf) gene has contributed to develop short-culm and high-yielding varieties widely grown in Southeast Asia and other areas. An incompletely dominant gene Ur1 (Undulate rachis -1) increases spikelet number per panicle, and can heighten grain yield in either the Ur1/Ur1 or Ur1/+ genotype by enlarging sink size. The effects of sd1 and Ur1 on lodging resistance and related traits were examined. The Ur1/Ur1, Ur1/+ and +/+ genotypes with sd1 (denoted by u, h and d, respectively, viz. three dwarfs) and those without sd1 (U, H and T, respectively, viz. three talls) under the common genetic background of a japonica variety Taichung 65 were grown in a field condition. sd1 reduced culm length by 29 to 37% and changed the internode elongation pattern toward the upper internode elongation type, whereas Ur1 increased culm length by 12 to 25% and changed toward the lower internode elongation type. The index of lodging was applied to the 4th internode, which was the 4th-panicle length (the total length from the 4th internode to panicle) × the 4th-top weight (the fresh weight including leaves at and above the 4th internode) / the breaking strength at the 4th internode. sd1 decreased the index of lodging by 47 to 59% with and without Ur1, being caused by the reduction of the 4th-panicle length and the increase of breaking strength, although its effect on reducing the 4th-top weight was not significant. The effect of sd1 on the breaking strength made a greater contribution towards the lodging-resistance enhancement than its height-reducing effect. Ur1, in the Ur1/Ur1 genotype, increased the index of lodging by 90 and 61% with and without sd1, respectively; while Ur1, in the Ur1/+ genotype, increased it by 73 and 32% with and without sd1. The increase of the index (deterioration of lodging resistance) by Ur1 was due to the reduction of breaking strength and the increases of both the 4th-panicle length and 4th-top weight. It is suggested that genes with lodging-resistance-enhancing effect should be introduced together with sd1 when Ur1 is utilized to develop high-yielding varieties.

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Heterosis is a heterozygous trait, depending on the genetic diversity of both parents. An approach to tag the heterosis trait with a molecular marker should be different from any other approaches for homozygous traits. To achieve this objective, it is necessary to identify the genetic diversity related to heterosis between both parents. This part of genetic diversity should be closely related to the parental combining ability, which is the most important factor for the performance of the hybrids. Based on the principle of parental combining ability, the simple sequence repeat (SSR) markers favorable and unfavorable to heterosis were identified in the previous study. In the present study, using these markers, two restorer lines with a very high combining ability, MGR44 and MGR45, were developed from the elite restorer line, Minhui63. In these two lines the favorable alleles were maintained while the unfavorable alleles were removed, resulting in the obvious increase of both the general combining ability (GCA) and special combining ability (SCA). Also, the spike density and 1000-grain weight traits were enhanced as compared with Minhui63. These results indicate that the previously selected molecular markers are very useful for hybrid rice breeding programs. Also, it was confirmed that the method described in our previous report is suitable and effective for screening molecular markers for hybrid rice breeding.

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Interspecific hybrids of Nicotiana repanda × N. tabacum die shortly after germination. Seedlings that are hybrids between N. repanda and N. tomentosiformis, which is a progenitor of N. tabacum, express hybrid lethality when they are cultured at 28°C for 90 days after germination, but seedlings that are hybrids between N. repanda and N. sylvestris, another progenitor of N. tabacum, grow normally. We detected several features of programmed cell death (PCD) in hybrid seedlings of N. repanda × N. tomentosiformis expressing hybrid lethality. Nuclear fragmentation was detected in nuclei isolated from dying shoots and roots of hybrid seedlings expressing lethality. In addition, electrophoresis of total DNA extracted from the shoots and roots of hybrid seedlings showed a distinctive DNA ladder pattern, suggesting cleavage of nuclear DNA into oligonucleosomal fragments. These results indicate that hybrid seedlings of N. repanda × N. tomentosiformis undergo PCD in the process of hybrid lethality. However, in hybrid seedlings of N. repanda × N. sylvestris, no features of PCD were detected. These results suggest that PCD might also be expressed in hybrid seedlings of N. repanda × N. tabacum because N. tomentosiformis is a progenitor of N. tabacum.

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A variant plant showing long internodes arose from a haploid plant obtained from clementine (Citrus clementina). Flow cytometric analysis and chromosome count revealed that it was a mixoploid with both haploid (2n = x = 9) and diploid (2n = 2x = 18) cells. The average length of the guard cells of the variant was almost the same as that of the haploid plant. This suggested that the epidermis of the variant consisted of haploid cells and that other leaf tissues consisted of diploid cells. Since in citrus the first germ layer forms the epidermis and the second and third layers form all the leaf tissues except for the epidermis, the variant was considered to be a ploidy periclinal chimera. When chromosomes of the young leaves were stained with chromomycin A₃ (CMA), they could be classified into four types based on the number and the position of the CMA-positive bands. The chromosome configuration of the variant differed from that of the original diploid clementine. The number of chromosomes of each type in the diploid cells of the variant was twice the number of those in the haploid plant. These results demonstrated that the diploid cells of the variant were doubled haploid cells, and that CMA analysis of chromosomes is useful for the identification of doubled haploids in citrus.

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Most commercial soybean varieties have yellow seeds due to loss of pigmentation in the seed coat. We previously showed that inhibition of seed coat pigmentation in yellow soybeans is controlled by post-transcriptional gene silencing (PTGS) of chalcone synthase (CHS) genes. Soybean CHS genes are composed of at least eight members, CHS1–CHS8. In this study, we compared the steady state mRNA level of each CHS member in the seed coat between a yellow soybean cultivar displaying CHS PTGS and its pigmented seed coat mutant in which the CHS PTGS is abrogated. Northern hybridization analysis suggested that in the pigmented seed coat, the transcripts of CHS7 and CHS8 were abundant among the total CHS transcripts, whereas those of other CHS members (CHS1–CHS6) were less abundant. Reverse transcription and subsequent real time PCRs using member-specific primers revealed reduction of the amount of mRNA of most CHS members in the non-pigmented seed coat of the yellow soybean as compared to the pigmented seed coat of the mutant, indicating the influence of PTGS on the CHS gene family.

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Since few genetic markers are available in bunching onion, many DNA markers are needed for the construction of a primary basis for the breeding of this crop. We report here the development of microsatellite markers in bunching onion. A size-fractionated genomic library was constructed from genomic DNA of bunching onion cv. ‘Kujo Futo’ and screened with a mixture of (GA)₁₅ and (GT)₁₅ oligonucleotide probes. From approximately 180,000 clones we isolated and sequenced 94 positive clones. A total of 52 clones were identified as having microsatellite repeats. Of them 49 had a GT motif, while only one had a GA motif, which is known to be more frequent than GT in most plant species. This is the first report of the abundance of GT over GA in Allium spp. Of 50 specific PCR primer pairs designed for the microsatellite-containing clones, 33 primer pairs amplified polymorphic loci in nine cultivars of bunching onion, 115 alleles detected in total. These results indicate that microsatellites with dinucleotide motifs, especially with GT, are promising sources of highly informative genetic markers in bunching onion.

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In higher plants, the genes related to the maintenance of the mitochondrial morphology are still largely unknown. In the present study, a rice dynamin-like protein, OsDRP3A, was identified as a rice homologue of Arabidopsis DRP3A and DRP3B proteins, which are involved in mitochondrial fission. Transient expression of a dominant-negative mutant of OsDRP3A containing the amino acid point mutation (K63A), caused mitochondria to become larger and less numerous than those in the native variety. A protein resulting from the fusion of green fluorescent protein (GFP) and wild-type OsDRP3A was observed, appearing in the form of punctate structures located at the tips and the constriction sites of mitochondria in live tobacco suspension-cultured (BY-2) cells. On the other hand, when GFP was fused to the dominant-negative mutant OsDRP3A (K63A), the mitochondrial morphology was distorted. These results suggest that OsDRP3A is involved in mitochondrial fission.

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A quantitative trait locus (QTL) analysis was carried out with a recombinant inbred line (RIL) population to identify the chromosomal regions responsible for cold tolerance of rice (Oryza sativa L.). The RIL population, consisting of 80 lines, was developed from a cross between the indica cultivar, Milyang 23 and the japonica weedy rice, Hapcheonaengmi 3. The population was genotyped with 2 morphological and 132 DNA markers, providing an average interval size of 11.3 cM, and was also evaluated for traits related to agricultural performance in cold water and in control plots. The RILs showed delayed heading and reduced culm length in the cold water plot and the differences in heading date and culm length between two plots were statistically significant. Cold tolerance was measured as days to heading, culm length, spikelet fertility, leaf discoloration, and panicle exsertion in the cold water plot, and difference in days to heading and the reduction ratio of culm length between two plots. A total of 14 QTLs for 7 traits were identified using single point and composite interval analysis. The number of QTLs per trait ranged from one to three. Phenotypic variation associated with each QTL ranged from 5.8 to 32.8%. No digenic interaction was detected. Several QTLs associated with cold tolerance were clustered in a few chromosomal blocks. For 11 (78.6%) of the QTLs identified in this study, the Hapcheonaengmi 3-derived alleles contributed desirable effects and favorable alleles were detected for difference in days to heading, spikelet fertility, panicle exsertion and leaf discoloration. From this study, it can be concluded that weedy rice is useful as a source of valuable alleles for breeding cold tolerance in rice.

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The presence of blast resistance genes in the elite Indica-type rice (Oryza sativa L.) varieties bred at the International Rice Research Institute (IRRI) was estimated based on a differential system using Philippines isolates of the rice blast fungus Pyricularia grisea (Cooke) Sacc., according to the gene-for-gene theory. Based on the presence of the three resistance genes, Pi20, Pita and one of the Pik alleles (other than Pik-s), the 42 varieties were classified into seven groups. A group that did not harbor these three genes contained seven varieties derived from the progenies of the hybrids with IR24 as a parental variety. The largest group harboring Pita consisted of 17 varieties, including IR36 and its sister lines or progenies. The group harboring by Pi20 had seven varieties that included IR8, IR24 and their hybrid progenies. Thus, most of the IRRI varieties were classified into these three groups that included IR8, IR24, IR36 or their hybrid progenies in their pedigree. The presence of a total of seven resistance genes Pib, Pita, Piz-t, Pi20, Pik-s, one of the Pik alleles (other than Pik-s) and one of the two genes, Pii or Pi3, was estimated in these varieties. In some cases, the presence of genes like Pib, Pik-s and Piz-t could not be confirmed due to the masking effect of Pita, Pik allele, or Pi20. The number and kind of blast resistance genes in IRRI varieties were limited compared with previously reported blast resistance genes. Since the presence of Pik-s and another Pik allele was estimated in 17 varieties belonging to five groups, and that of Pib in 38 varieties belonging to four groups, it appeared that these genes were widely distributed in IRRI-bred varieties.

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Optimizing hybrid vigor (HV) has been a primary objective in hybrid crop breeding programs. The effective use of DNA markers in this endeavor has been plagued by inconsistent results among researchers who investigated the relationship between genetic similarity (GS) among parents and heterosis in their F₁’s. The objectives of this study were to evaluate the relationship between GS among parents and heterosis in F₁ for grain yield in japonica rice, and to formulate a strategy for marker-based prediction of heterosis. Forty-five F₁ hybrids were generated from a half-diallel design using 10 japonica rice cultivars. Parents and hybrids were grown under medium N and 0-N fertilizers and evaluated for grain yield. The parental varieties were genotyped with 188 SSR markers and/or 129 RAPD primers. Estimates of GS based on all markers were not highly correlated with heterosis. After analyzing the association of each marker to yield heterosis, we were able to identify markers that were significantly associated with HV in this collection of japonica rices. The estimates of GS based on this set of selected markers showed highly significant correlations with HV. In addition, some SSR and RAPD markers were found to be associated with hybrid weakness (HW). This suggests that the heterosis observed in the F₁’s could be improved by selectively combining alleles associated with HV and eliminating those associated with HW. A new concept for developing “key markers” is proposed and the feasibility of predicting heterosis through estimation of GS among parents utilizing key DNA markers is discussed.

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Soybean (Glycine max (L.) Merrill) is the most important leguminous crop in the world due to the high contents of protein and oil, and accumulation of various physiologically active substances. However, most of the genomic and economic traits in soybean are quantitative, controlled by multiple genes and easily affected by environmental conditions. Based on recombinant inbred lines (F₈), a genetic linkage map with 177 RFLP, 150 SSR, 28 AFLP markers and 5 phenotypic markers was constructed. The map covered a distance of 2663.6 cM of the soybean genome comprising 20 linkage groups. The average distance between two adjacent marker loci was 7.89 cM. In this population, we detected thirty-nine QTLs for all the reproductive development and seed quality traits investigated, that is, three for flowering time (FT1–3), four for maturity (HAV1–4), three for reproductive period (RP1–3), three for seed hardness (RAS1–3), five for viability of seed (VIS1–5), four for germination rate of seed (GRS1–4), five for water absorbability of seed (WAS1–5) and twelve QTLs for seed weight (SWE1–6 and SWH1–6). Out of these QTLs, twenty-eight were detected in nearly the same regions of the linkage map by both IM (interval mapping) and CIM (composite interval mapping) analysis. The proportion of variance explained of these QTLs ranged from 3.4% to 67.1%. Epistatic interactions were detected among various QTLs. Especially there was a strong interaction among the effects of FT1 and FT2, and FT1 and FT3. Multiple correlation coefficients between FT1 and FT2, and FT1 and FT3 accounted for 79.6% and 74.1% of the phenotypic variation of flowering time, respectively.

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The uzu gene shows typical semi-dwarf plant type and has resulted in lodging resistance and improved canopy structure in barley (Hordeum vulgare L.). The geographic distribution of ‘uzu’ lines is limited in East Asia including Japan, Korean peninsula and China. For many years, majority of cultivated six-rowed barley varieties in southern Japan were ‘uzu’ type. Inheritance of the uzu gene, located on chromosome 3H, is monogenic recessive and the expression is pleiotropic during developmental process. In cereal crop plants, several semi-dwarf genes have been widely used in breeding programs to increase productivity and it is known that several of these semi-dwarf genes were derived from gibberellin (GA)-related mutations. The barley dwarfing gene uzu is independent to GA. This characteristics are known earlier. In this study, we characterized uzu gene in molecular aspects. As the result of comprehensive study using several pairs of isogenic lines, we found the morphological and physiological similarities of barley uzu gene to rice dwarf mutant, d61. D61 encodes OsBRI1, which is homologous gene for Brassinosteroid insensitive 1 in Arabidopsis. We also found that ‘uzu’ lines specific single nucleotide polymorphism (SNP) contributed for amino acid substitution in barley homologous sequence of OsBRI1, HvBRI1. Linkage analysis using a segregating population for uzu showed the co-segregation between uzu and HvBRI1. Furthermore, as the results of derived cleaved amplified polymorphic sequences (dCAPS) marker analysis using more than 260 ‘uzu’ landraces and the genomic sequencing analysis of HvBRI1 gene derived from 19 barley accessions, all the ‘uzu’ lines investigated in this study had the same SNP in putative kinase domain of HvBRI1. On the basis of these findings, we discussed the phylogeny of ‘uzu’ landraces in East Asia.

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