宇野 雄一 | ![]() |
ウノ ユウイチ | |
大学院農学研究科 資源生命科学専攻 | |
教授 | |
農学関係 |
[査読有り][招待有り]
研究論文(学術雑誌)
研究論文(学術雑誌)
[査読有り]
研究論文(学術雑誌)
The strawberry fruit contains abundant polyphenols, such as anthocyanins, flavan-3-ol, and ellagitannin. Polyphenol enrichment improves the quality of strawberries and leads to a better understanding of the polyphenol induction process. We measured the total polyphenol content of strawberry fruits under different growth conditions, developmental stages, and treatment conditions during pre-harvest and post-harvest periods. High fruit polyphenol content was observed in cold treatment, which was selected for further analysis and optimization. A transcriptome analysis of cold-treated fruits suggested that the candidate components of polyphenols may exist in the phenylpropanoid pathway. Coverage with a porous film bag excluded the effects of drought stress and produced polyphenol-rich strawberry fruits without affecting quality or quantity. The degree of stress was assessed using known stress indicators. A rapid accumulation of abscisic acid was followed by an increase in superoxide dismutase and DPPH (2,2-Diphenyl-1-picrylhydrazyl) activity, suggesting that the strawberry fruits responded to cold stress immediately, reaching the climax at around 6 days, a trend consistent with that of polyphenol content. These findings enhance our understanding of the mechanism of post-harvest polyphenol accumulation and the value of strawberries as a functional food.
MDPI AG, 2022年, Plants, 11 (17), 17 (13 pages) - 2220, 英語, 国際誌[査読有り][招待有り]
研究論文(学術雑誌)
Rab proteins are small GTP-binding proteins and are the largest family in the Ras GTPase superfamily and mediate vesicular transport in cells. Diverse insulin-like peptides, such as bombyxin, are synthesized in the brain and secreted into the haemolymph by the corpus allatum (CA). In the brain of Bombyx mori, Rabs are expressed in a specific area; however, which Rabs actually link the secretion of bombyxin remains unknown. A double-staining analysis of nine Rabs ( Rab1, 3, 6, 7, 14, 21, 26, 39 and X4) and bombyxin indicated that Rab3-, Rab7-, Rab39-and RabX4-immunohistochemical reactivity (ir) areas overlapped with bombyxin-ir in the brain and CA in B. mori, while Rab6-, Rab14-and Rab21-irs partially overlapped in the CA. Rab1-ir occurred in the other immunopositive areas in CA. Rab26-ir did not occur in the brain. Rab39-ir occurred in UNC104, Rab39- effector,-immunopositive neurons in the brain and CA. Thus, Rab3, 7, 39 and X4 may regulate the exocytosis of bombyxin.
CZECH ACAD SCI, INST ENTOMOLOGY, 2021年, EUROPEAN JOURNAL OF ENTOMOLOGY, 118, 307 - 314, 英語[査読有り]
研究論文(学術雑誌)
Efficient cultivation methods were investigated to promote the branding of products in plant factories. Moderate stress can enhance plant constituents that are beneficial for human health, without reducing yield. Dehydration stress in lettuce rhizospheres increased some antioxidants, including L-ascorbic acid (AsA) and polyphenols. In this study, the major factors contributing to the augmentation of antioxidant constituents were investigated. The drought treatment resulted in increased hydrophilic oxygen radical absorbance capacity (ORAC) values but not hydrophobic ORAC values. Both activities of antioxidant enzymes (superoxide dismutase, SOD, and ascorbate peroxidase, APX) were elevated under drought conditions. RNA-seq analysis revealed 33 upregulated and 115 downregulated differentially expressed genes, and 40 gene ontology enrichment categories. A dehydrin gene was the most significant among the upregulated genes in response to drought stress. Dehydrin protects plant cells from dryness through multiple functions, such as radical scavenging and protection of enzymes. Real-time PCR validated the substantial increase in some dehydrin paralogs with root desiccation. In conclusion, the enhancement of antioxidant levels by drought stress is likely not due to the induction of antioxidant enzyme genes, but due to increased enzymic activities. These activities might be protected by dehydrins encoded by the upregulated paralogs under drought stress.
MDPI, 2021年, Horticulturae, 7 (11), 444, 英語[査読有り]
研究論文(学術雑誌)
African violet (Saintpaulia ionantha) is an herbaceous perennial of the Gesneriaceae family. Because almost all the cultivars are heterozygous, pure lines are useful for both classical and new breeding approaches. A shortcut to obtain purebred lines involves the production of doubled haploid strains produced from anther-derived haploids. In this chapter, a protocol for culturing African violet anthers is described in detail.
2021年, Methods in molecular biology (Clifton, N.J.), 2289, 249 - 261, 英語, 国際誌[査読有り]
研究論文(学術雑誌)
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研究論文(学術雑誌)
[査読有り]
研究論文(学術雑誌)
[査読有り]
研究論文(学術雑誌)
[査読有り]
研究論文(学術雑誌)
[査読有り]
研究論文(学術雑誌)
[査読有り]
研究論文(学術雑誌)
Fra a 1 is a strawberry allergen that causes oral allergic syndrome. Fra a 1.01 is a major isoform that accumulates abundantly in fruits during the winter season. Here, we tested the hypothesis that Fra a 1.01 responds to environmental factors, such as cold stress. We analyzed transcriptional and translational levels of Fra a 1.01 in strawberry calli and organs under various cold conditions. First, we incubated strawberry (Fragaria x ananassa 'Akihime') calli and post-harvested fruits at low temperatures from several hours to days. Fra a 1.01 did not show significant differences in either gene expression or protein accumulation levels, suggesting that short-term cold treatments did not affect Fra a 1.01 expression. Second, we exposed whole plants to low temperature conditions for similar to 28 days. Under conditions below 10 degrees C, Fra a 1.01 transcripts were induced gradually throughout the cold treatment (crown and root), or from 2 days to the last day (leaf and fruit). The Fra a 1.01 protein remarkably accumulated in crowns and slightly in fruits after 28 days. Finally, the promoter region of Fra a 1.01e was analyzed to detect tissue-specific expression. The cloned and sequenced promoter included several cis-acting regulatory elements related to cold response. When the Fra a 1.01 promoter region was heterologously expressed in Arabidopsis, the promoter activities, as assessed by GUS staining, were observed mainly around the shoot apices and in roots. Thus, Fra a 1.01 was considered to be expressed in crowns and roots, and was additively induced by cold stress. These organ-specific expressions could be important in elucidating the mechanisms responsible for Fra a 1.01 protein's accumulation in fruits during the winter season.
JAPAN SOC HORTICULTURAL SCI, 2020年, HORTICULTURE JOURNAL, 89 (2), 182 - 190, 英語[査読有り]
研究論文(学術雑誌)
To improve several agronomic traits in crisphead lettuce (Lactuca sativa L.) under high-temperature growth conditions, we investigated the correlation among those traits in multiple cultivars and performed genetic mapping of their causal genes. In a field cultivation test of Empire type (serrated leaf) and Salinas type (wavy leaf) cultivars, Empire type cultivars showed increased tipburn susceptibility and late bolting compared with Salinas type cultivars. We attempted genetic mapping of leaf shape and bolting time by ddRAD-seq using the F2 population derived from a cross between 'VI185' (Empire type) and 'ShinanoGreen' (Salinas type). These analyses suggested that both traits are controlled by a single locus in LG5. Genotyping of 51 commercial lettuce cultivars with a tightly linked marker (LG5_v8_252.743Mbp) at this locus showed an association between its genotype and the serrated leaf phenotype. By further fine mapping and transcriptome analysis, a gene encoding putative CIN-like TCP transcription factor was determined to be a candidate gene at this locus and was designated as LsTCP4. An insertion of retrotransposable element was found in the allele of 'VI185', and its transcript level in the leaves was lower than that in 'ShinanoGreen'. Because shapes of leaf epidermal cells in 'VI185' were similar to those in the TCP family mutant of Arabidopsis thaliana, the leaf shape phenotype was likely caused by reduced expression of LsTCP4. Furthermore, because it is known that the TCP family protein also controls flowering time via interaction with FT in A. thaliana, it was highly possible that LsTCP4 gave pleiotropic effects on both leaf shape and bolting time in lettuce.
2020年, Horticulture research, 7, 15 - 15, 英語, 国際誌[査読有り]
研究論文(学術雑誌)
© 2019 Elsevier B.V. Flowering time is one of the most important traits in carnation (Dianthus caryophyllus L.) breeding because it decides the yearly yield. Two previously developed mapping populations were used to determine the number and effect of quantitative trait loci (QTLs) for flowering time. Flowering time showed large phenotypic segregation in two F2 populations and in different years. Despite the different populations and different years, one major common QTL for flowering time was detected in linkage group 10 with the effect explaining from 18.2%–22.5% of the overall phenotypic variance. We developed a DNA marker, qD1Flw1-sc43-4, that was located close to the detected QTL for flowering time. We could distinguish the flowering time and categorize the genotypes of an F1 population derived from a cross between late flowering ‘Light Pink Barbara’ and early flowering ‘Kaneainou 1 go’ using the qD1Flw1-sc43-4 marker. Our results suggest that flowering time in carnation involves several genetic factors. In this study, we identified one of the major factors for flowering time and developed a DNA marker that was tightly linked to the major QTL.
ELSEVIER, 2020年, Scientia Horticulturae, 262, 109053, 英語[査読有り]
研究論文(学術雑誌)
Sufficient yields of high-quality RNA are needed for next-generation sequencing and high-throughput real-time polymerase chain reaction analyses. In the case of strawberry (Fragaria x ananassa) fruits, successful RNA isolation requires removal of abundant inhibitory substances (polysaccharides and polyphenols) that greatly reduce quality and yield. In this study, we applied various combinations of RNA isolation protocols directed at reproductive organs. The best manual isolation method involved nonionic polymer and modified acid guanidinium thiocyanate-phenol-chloroform treatments followed by phenol/chloroform/isoamyl alcohol extraction. Compared with other methods, this approach gave significantly higher yields [84.0 mu g/g fresh weight (FW)] of RNA of greater purity (A260/A280 = 1.99; A260/230 = 1.51). Better-quality RNA (A260/230 = 2.11) was obtained using an automated method, but the yield was lower (18.1 mu g/g FW) than that obtained manually. This automated method consisted of pretreatment with nonionic polymer followed by a silica-based system extraction. Although RNA of sufficient quality [RNA Integrity Number (RIN) >= 6.5 and 28S/18S >= 1.0] for RNA sequencing was obtained from receptacles using both automated and manual methods, the manual method yielded high-quality RNA from achenes and anthers. The automatic method features 6-fold faster high-throughput capacity, whereas the manual method has wider applicability to different tissues.
AMER SOC HORTICULTURAL SCIENCE, 2019年02月, HORTSCIENCE, 54 (2), 380 - 384, 英語[査読有り]
研究論文(学術雑誌)
[査読有り]
研究論文(学術雑誌)
Strawberry (Fragaria x ananassa) contains a major allergen, Fra a 1, which causes oral allergic syndrome. Fra a 1 is a PR-10 homolog that is regulated by environmental conditions. The allergenicity of fruit caused by Fra a 1 may depend on the genotype or growing conditions. We analyzed the Fra a 1.01 transcript levels and Fra a 1.01 protein levels in strawberry fruits of several genotypes across all seasons. In the preliminary rough screening, we selected the line WH1 bearing white fruit and the red-fruited cultivar 'Akihime'. Under the same environmental conditions, there was no significant difference in Fra a 1.01 levels between the two cultivars over several months, suggesting that receptacle color was not indicative of allergenicity caused by Fra a 1.01. Fruits cultivated under the same environmental conditions should be used for comparisons of the allergenicity among genotypes. Both 'Akihime' and WM accumulated significantly higher levels of Fra a 1.01 protein in winter than in spring. We investigated the effects of irradiation and low temperature as environmental factors controlling the accumulation of Fra a 1.01 in winter. A shading treatment on fruit did not significantly affect Fra a 1.01 protein accumulation in straw berry fruits. Regarding variations over time, the Fra a 1.01 protein content was higher in fruits harvested at midnight in January than in those harvested at other times and in other months. These findings suggested that the Fra a 1.01 protein accumulates in response to environmental factors such as cold stress.
JAPAN SOC HORTICULTURAL SCI, 2019年, HORTICULTURE JOURNAL, 88 (3), 354 - 363, 英語[査読有り]
研究論文(学術雑誌)
CYP2C9 is a human microsomal cytochrome P450c (CYP). Much variation in CYP2C9 levels and activity can be attributed to polymorphisms of this gene. Wild-type CYP2C9 and ten mutants were co-expressed with NADPH-cytochrome P450 reductase in Escherichia coli. The hydroxylase activities toward steroids were examined. CYP2C9.2, CYP2C9.3, CYP2C9.4, CYP2C9.16, CYP2C9.28, CYP2C9.48 and
2018年08月, Biopharmaceutics & Drug Disposition, 39 (8), 371 - 377, 英語[査読有り]
研究論文(学術雑誌)
Fra a 1 protein in strawberry causes oral allergic syndrome. Over 39 Fra a 1 paralogs have been identified in strawberry genome. Fra a 1.01 is major accumulating protein in edible organs.Strawberry fruits contain allergenic proteins that cause oral allergic syndrome. The hypothesized major allergen is Fra a 1, an ortholog of the birch pollen allergen protein Bet v 1. We organized Fra a 1 genes and analyzed their localizations at the transcriptional and translational levels. In total, 15 new Fra a 1 proteins were identified from the genomic database, increasing the total number of Fra a 1 to 30 proteins encoded by 39 genes. Fra a 1.02 was mostly expressed in receptacles, and Fra a 1.01 in achenes, when analyzed by RNA sequencing. Immunoblotting showed that the Fra a 1.01 protein was broadly accumulated in strawberry organs, while the Fra a 1.02 protein was mostly expressed in receptacles. Recombinant Fra a 1.01 strongly reacted with human IgE. The mRNA and protein expression levels of Fra a 1 did not correlate, indicating the importance of protein levels when evaluating the abundance of allergens in strawberry. Based on the localizations, accumulation levels and reactivity to human IgE, we determined that Fra a 1.01 was the most important allergen, followed by Fra a 1.02, and then other Fra a 1 proteins. The information obtained here will be useful for selecting the target Fra a 1 paralogs when breeding hypoallergenic strawberry.
Springer, 2018年03月, Plant Cell Reports, 37 (3), 411 - 424, 英語[査読有り]
研究論文(学術雑誌)
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研究論文(学術雑誌)
This study investigated the effect of the red (R) to far-red (FR) photon flux density ratio on the growth and crocin yields of 64 saffron (Crocus sativus L.) corms that were hydroponically cultivated in closed chambers. The life cycle of saffron can be divided into four stages: formation of the flower buds, flowering, formation of the daughter corms (FD) and development of the daughter corms (DD). During the DD stage, the saffron corms were cultivated under the same environmental conditions except for the light quality, which was applied at an R/FR ratio of 15.8 (FL treatment) or 1.8 (FR treatment). There was no significant difference between treatments in the shoot length, or the maximum diameter, weight and stigma weight of the daughter corms. However, there was a significant difference in the absorbance of crocin solutions. Although the cause of FR-induced increase in crocin was not elucidated, it was presumed that a low R/FR ratio during the DD stage accelerates the translocation of photosynthetic products from the leaves to the corms to generate carbohydrate-enriched corms. This might result from increased sink strength, which is associated with phytochrome equilibrium.
Biotron Institute, 2018年, Environmental Control in Biology, 56 (2), 51 - 57, 英語[査読有り]
研究論文(学術雑誌)
The aim of this study was to clarify how the air temperatures during the light and dark periods affect growth and biosynthesis of the iridoid glycoside asperuloside in Hedyotis diffusa. Ten experimental air temperature treatments were established in a growth chamber during the cultivation period: (light/dark period air temperature) 30/15°C, 20/20°C, 25/20°C, 30/20°C, 35/20°C, 25/25°C, 30/25°C, 35/25°C, 40/25°C and 30/30°C. It was found that the stem length and epigeous dry weight were greatest with the 35/25°C treatment, whereas the leaves near the apical buds of the branches became blackened with the 40/25°C treatment, indicating that 35°C represents the upper air temperature limit during the light period for growth without any physiological disorder. By contrast, the asperuloside concentration was high with an air temperature of 25°C or less during the light period and 20°C or less during the dark period, and was greatest with the 25/20°C and 30/20°C treatments. Thus, air temperatures of 2530°C during the light period and 20°C during the dark period are effective for maximising the asperuloside content in H. diffusa.
Biotron Institute, 2018年, Environmental Control in Biology, 56 (2), 73 - 79, 英語[査読有り]
研究論文(学術雑誌)
CYP2C9 is a human microsomal cytochrome P450c (CYP). Much of the variation in CYP2C9 levels and activity can be attributed to polymorphisms of this gene. Wild-type CYP2C9 and mutants were coexpressed with NADPH-cytochrome P450 reductase in Escherichia coli. The hydroxylase activities toward 7-ethoxycoumarin, flavanone and steroids were examined. Six CYP2C9 variants showed Soret peaks (450nm) typical of P450 in reduced CO-difference spectra. CYP2C9.38 had the highest 7-ethoxycoumarin de-ethylase activity. All the CYP2C9 variants showed lower flavanone 6-hydroxylation activities than CYP2C9.1 (the wild-type). CYP2C9.38 showed higher activities in testosterone 6-hydroxylation, progesterone 6-/16-hydroxylation, estrone 11-hydroxylation and estradiol 6-hydroxylation than CYP2C9.1. CYP2C9.40 showed higher testosterone 17-oxidase activity than CYP2C9.1; CYP2C9.8 showed higher estrone 16-hydroxylase activity and CYP2C9.12 showed higher estrone 11-hydroxylase activity. CYP2C9.9 and CYP2C9.10 showed similar activities to CYP2C9.1. These results indicate that the substrate specificity of CYP2C9.9 and CYP2C9.10 was not changed, but CYP2C9.8, CYP2C9.12 and CYP2C9.40 showed different substrate specificity toward steroids compared with CYP2C9.1; and especially CYP2C9.38 displayed diverse substrate specificities towards 7-ethoxycoumarin and steroids.
WILEY, 2017年11月, BIOPHARMACEUTICS & DRUG DISPOSITION, 38 (8), 486 - 493, 英語[査読有り]
研究論文(学術雑誌)
We investigated the growth characteristics and physiological activity of leaf lettuce cultivated using different sizes of liquid fertilizer droplets and different rates of fog flow through the rhizosphere using a dry-fog hydroponics technique. These characteristics were subsequently compared to lettuce cultivated using deep flow technique (DFT), which is the prevailing hydroponic technique used in greenhouses. The cultivation chambers were filled with dry-fog (DF) containing droplets that average less than 10 μm in diameter or semi-dry-fog (SDF) containing droplets that average 20 μm in diameter. The plants were grown for three weeks following transplantation, and we measured the growth, respiration rate and TTC reduction ability of the roots in addition to the photosynthesis rate and the contents of total soluble protein, nitrate nitrogen and calcium ions. The amount of leaf growth increased with a slower flow rate under both DF and SDF conditions and was increased in both dry-fog aeroponic conditions compared to leaves grown in DFT. SDF with a slow flow rate increased root growth and activity and resulted in an increase in stomatal conductance and photosynthesis rate. DF with a slow flow rate also significantly increased the growth of leaves and roots, but the physiological activities were not affected. Thus, the growth and physiological characteristics of leaf lettuce cultivated using dry-fog aeroponics was improved in different ways by changing both the atomized droplet size and flow rate in the root zone.
International Society for Horticultural Science, 2017年10月30日, Acta Horticulturae, 1176, 45 - 51, 英語[査読有り]
研究論文(学術雑誌)
A new spray fertigation system (dry-fog hydroponics) was developed for establishing ever-bearing strawberry (Fragaria × ananassa Duch. 'Summer ruby') production during the summer. Dry-fog hydroponics is an aeroponic technique that sprays a very fine foggy nutrient solution with an average droplet diameter of less than 10 μm into the root zone. The roots extend into the chamber filled with dry-fog of the liquid fertilizer and absorb water and nutrients from the dry-fog that fills the rhizosphere. This aeroponic system needs less water than any other hydroponic technique and keeps rhizosphere temperature lower than air temperature owing to the release of latent heat by dry-fog evaporation and adiabatic expansion of solution from the exhaust nozzle. Runner seedlings of strawberry plants were cultivated with dry-fog aeroponic system from early June to December in a greenhouse, and the growth, flower bud formation, fruit quality, photosynthetic rates as well as the contents of chlorophyll and total soluble protein in leaves were compared with those of plants cultivated by drip fertigation with palm shell medium as a control. Rhizosphere and the crown temperatures of dry-fog aeroponics were reduced by approximately 3-5°C throughout the day compared with control and were maintained below 25°C during the late summer. Although ever-bearing strawberry plants are insensitive to high temperature under long day conditions from a viewpoint of flower initiation, low temperature, especially the crown part of shoot base, is considered important for inducing flowering. For dry-fog aeroponics, the number of inflorescences and °Brix of fruits significantly increased but the number of runners and vegetative growth significantly decreased compared with control. Photosynthetic rates of leaves grown with dry-fog aeroponics increased compared to those of the control under both light and CO2 limited and saturated measurement conditions. In addition, leaf chlorophyll and total soluble protein contents significantly increased in dry-fog aeroponics compared with the control. From these findings, we suggested that dry-fog aeroponics is a usable new hydroponics for raising ever-bearing strawberry plants during the summer by maintaining rhizosphere temperature low, and thus suitable for shifting their growth phase from vegetative to reproductive.
International Society for Horticultural Science, 2017年10月30日, Acta Horticulturae, 1176, 37 - 44, 英語[査読有り]
研究論文(学術雑誌)
Rab proteins are small monomeric GTPases/GTP-binding proteins, which form the largest branch of the Ras superfamily. The different Rab GTPases are localized to the cytosolic face of specific intracellular membranes, where they function as regulators of distinct steps in membrane trafficking. RabX4 is an insect-specific Rab protein that has no close homolog in vertebrates. There is little information about insect-specific Rab proteins. RabX4 was expressed in Escherichia coli and subsequently purified. Antibodies against Bombyx mori RabX4 were produced in rabbits for western immunoblotting and immunohistochemistry. Western blotting of neural tissues revealed a single band, at approximately 26kD. RabX4-like immunohistochemical reactivity was restricted to neurons of the pars intercerebralis and dorsolateral protocerebrum in the brain. Further immunohistochemical analysis revealed that RabX4 colocalized with Rab6 and bombyxin in the corpus allatum, a neuronal organ that secretes neuropeptides synthesized in the brain into the hemolymph. RabX4 expression in the frontal ganglion, part of the insect stomatogastric nervous system that is found in most insect orders, was restricted to two neurons on the outer region and did not colocalize with allatotropin or Rab6. Furthermore, RNA interference of RabX4 decreased bombyxin expression levels in the brain. These findings suggest that RabX4 is involved in the neurosecretion of a secretory organ in Bombyx mori.
WILEY, 2017年09月, ARCHIVES OF INSECT BIOCHEMISTRY AND PHYSIOLOGY, 96 (1), 英語[査読有り]
研究論文(学術雑誌)
Strawberry fruit contains many constituents, some of which have the potential to inhibit histidine decarboxylase (HDC) activity. HDC converts L-histidine to histamine, which is associated with allergic and other biological reactions in the human body. The HDC inhibition levels were different and the component ratios varied by genotype in strawberry. Among the 11 cultivars collected locally in Japan, 'Tokun' had an approximately ten times higher inhibition ratio than the lowest cultivar. The reproducibility was confirmed using five cultivars under the same conditions in a glass greenhouse, suggesting that genotypic variation is a major factor of HDC inhibition. The potential inhibitors of HDC might be polyphenols because they showed moderate correlations with HDC inhibition rates. Among the polyphenols, the anthocyanin content possessed a moderate negative correlation. Ascorbic acid, which contributes to the overestimation of total polyphenol, did not independently inhibit HDC activity. These findings will support the identification of potential HDC inhibitors in strawberry and indicated that genotypic differences would make useful probes for inhibitor identification.
SPRINGER HEIDELBERG, 2017年06月, ACTA PHYSIOLOGIAE PLANTARUM, 39 (6), 6 pages, 英語[査読有り]
研究論文(学術雑誌)
Strawberry fruit contain the allergenic Fra a proteins, members of the pathogenesis-related 10 protein family that causes oral allergic syndrome symptoms. Fra a proteins are involved in the flavonoid biosynthesis pathway, which might be important for color development in fruits. Auxin is an important plant hormone in strawberry fruit that controls fruit fleshiness and ripening. In this study, we treated strawberry fruits with exogenous auxin or auxin inhibitors at pre- and post-harvest stages, and analyzed Fra a transcriptional and translational expression levels during fruit development by real-time PCR and immunoblotting. Pre-harvest treatment with 1-naphthaleneacetic acid (NAA) alone did not affect Fra a expression, but applied in conjunction with achene removal NAA promoted fruit pigmentation and Fra a protein accumulation. The response was developmental stage-specific: Fra a 1 was highly expressed in immature fruit, whereas Fra a 2 was expressed in young to ripe fruit. In post-harvest treatments, auxin did not contribute to Fra a induction. Auxin inhibitors delayed fruit ripening; as a result, they seemed to influence Fra a 1 expression. Thus, Fra a expression was not directly regulated by auxin, but might be associated with the ripening process and/or external factors in a paralog-specific manner.
MDPI, 2017年06月, INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES, 18 (6), 1186 (12 pages), 英語[査読有り]
研究論文(学術雑誌)
Tipburn is a physiological disorder caused by a calcium (Ca) deficiency that occurs mainly in leafy vegetables, such as lettuce, resulting in a reduced commercial value. To prevent tipburn injury, a sensitive cultivar was tested as an indicator for early symptom detection. An indicator cultivar began to develop tipburn two days earlier than the target cultivar. This allowed for the rescue of the target cultivar by the addition of extra Ca. The yield rate was improved from 4% to 70% with the use of an indicator cultivar in the hydroponic cultures. The top fresh weight of the target cultivar when using an indicator cultivar decreased compared with control plants, but increased compared with negative control plants lacking Ca in the culture. Water contents and root lengths were not affected by the use of an indicator cultivar and additional Ca. These results were consistent with other target cultivars under excess ammonium conditions. This system may be used against tipburn incidence without additional costs and equipment in plant factories. (C) 2016 Elsevier B.V. All rights reserved.
ELSEVIER SCIENCE BV, 2016年10月, SCIENTIA HORTICULTURAE, 210, 14 - 18, 英語[査読有り]
研究論文(学術雑誌)
In eukaryotic cells, Rab guanosine triphosphate-ases serve as key regulators of membrane-trafficking events, such as exocytosis and endocytosis. Rab3, Rab6, and Rab27 control the regulatory secretory pathway of neuropeptides and neurotransmitters. The cDNAs of Rab3, Rab6, and Rab27 from B. mori were inserted into a plasmid, transformed into Escherichia coli, and then subsequently purified. We then produced antibodies against Rab3, Rab6, and Rab27 of Bombyx mori in rabbits and rats for use in western immunoblotting and immunohistochemistry. Western immunoblotting of brain tissue revealed a single band at approximately 26 kDa. Immunohistochemistry results revealed that Rab3, Rab6, and Rab27 expression was restricted to neurons in the pars intercerebralis and dorsolateral protocerebrum of the brain. Rab3 and Rab6 co-localized with bombyxin, an insect neuropeptide. However, there was no Rab that co-localized with prothoracicotropic hormone. The corpus allatum secretes neuropeptides synthesized in the brain into the hemolymph. Results showed that Rab3 and Rab6 co-localized with bombyxin in the corpus allatum. These findings suggest that Rab3 and Rab6 are involved in neurosecretion in B. mori. This study is the first to report a possible relationship between Rab and neurosecretion in the insect corpus allatum.
SPRINGER, 2016年07月, HISTOCHEMISTRY AND CELL BIOLOGY, 146 (1), 59 - 69, 英語[査読有り]
研究論文(学術雑誌)
To optimize the production of anther-derived haploid plants of the genus Saintpaulia, the effect of plant growth regulator (PGR) concentration in the induction medium was investigated for six Saintpaulia species and eight cultivars of S. ionantha. The most effective PGR concentration for shoot formation was 1.0 mg l(-1) for both N-6-benzyladenine and alpha-naphthaleneacetic acid among the nine combinations of three concentrations (0.1, 1.0 and 10.0 mg l(-1)) of each PGR. Microspores at the uninucleate stage, which is the most suitable stage for anther culture, were observed in buds 2-3 mm (in Saintpaulia species) or 3-5 mm (in S. ionantha cultivars) in length. The frequency of shoot formation from anther-derived calli ranged from 0 to 63 % for Saintpaulia species and 0 to 100 % for S. ionantha cultivars. Five Saintpaulia species and seven S. ionantha cultivars formed shoots under the same PGR concentrations. These results demonstrated the versatility of the optimized conditions for anther cultures of diverse genotypes in Saintpaulia. Microscopic observation of the chromosomes of root-tip cells of anther-derived plants showed that the haploid efficiency was 40.0-85.7 % (average 66.6 %). Some haploid individuals showed phenotypic differences from the diploid parents in flower color and shape. Significant differences in stomatal guard cell lengths were observed between the diploid parent and some of the anther-derived haploids. The described protocol enabled haploids of Saintpaulia species to be obtained in a minimum of 286 days.
SPRINGER, 2016年05月, PLANT CELL TISSUE AND ORGAN CULTURE, 125 (2), 241 - 248, 英語[査読有り]
研究論文(学術雑誌)
研究論文(国際会議プロシーディングス)
研究論文(国際会議プロシーディングス)
研究論文(国際会議プロシーディングス)
研究論文(国際会議プロシーディングス)
研究論文(国際会議プロシーディングス)
The purpose of this study was to clarify the effects of light intensity and light/dark period on the concentration of iridoids in the plant Hedyotis diffusa, which finds use in herbal medicine. To investigate the effect of light conditions, plants were grown in a chamber with factorial treatments comprising three light/dark periods (14/10 h, 19/5 h and 24/0 h) and two light intensities (photosynthetic photon flux density of 142 and 40μmol m-2 s-1). The epigeous dry weight was greatest with the treatments using high light intensity regardless of light period. The concentration of asperuloside (an iridoid) at 24/0 h was noticeably greater than that at 14/10 h or 19/5 h irrespective of light intensity. It was expected that the critical light period for the production of asperuloside would occur between 19 h and 24 h. The content of asperuloside was greatest at 24/0 h under high light intensity. Therefore, high light intensity and a 24/0-h light/dark period can produce high-quality H diffusa.
Biotron Institute, 2016年, Environmental Control in Biology, 54 (2), 109 - 116, 英語[査読有り]
研究論文(学術雑誌)
The growth characteristics and physiological activities of leaves and roots of lettuce cultivated in dry-fog aeroponics with different flow rates of nutrient dry-fog (FL, 1.0 m s-1 NF, 0.1 m s-1) were investigated under a controlled environment for two weeks and compared to lettuce cultivated using deep-flow technique (DFT). The growth of leaves of FL and DFT was not different and was significantly higher than that of NF. The amount of dry-fog particles adhering to the objects was higher in FL than in NF, so that the root growth in NF was significantly higher than that of FL. The respiration rate of roots was significantly higher in dry-fog aeroponics, but the dehydrogenase activity in the roots was significantly higher in DFT. There were no differences in the contents of chlorophyll and total soluble protein in the leaves or the specific leaf area. Photosynthetic rate and stomatal conductance were higher in dry-fog aeroponics. The contents of nitrate nitrogen, phosphate and potassium ions in the leaves were significantly higher in DFT, but the content of calcium ions was significantly higher in FL. Thus, changing the flow rate of the dry-fog in the rhizosphere can affect the growth and physiological activities of leaves and roots.
Biotron Institute, 2015年, Environmental Control in Biology, 53 (4), 181 - 187, 英語[査読有り]
研究論文(学術雑誌)
The purpose of this study is to measure the time variation of nitrate concentration in whole lettuce plants throughout cultivation. Lettuce plants were cultivated in growth chambers under fluorescent or light-emitting diode (LED) lamps. Four different light/dark periods and two ratios of red to blue photon flux density (R/B ratio) were prepared. The cultivation terms utilized were 1 or 2 weeks, and 289 lettuce plants were harvested. The visible-near-infrared absorption spectrum and nitrate concentration were obtained for each sample by spectrometry and the RQflex method, respectively. A regression model for the nitrate concentration was constructed from the spectra, and the correlation coefficient and standard error between the measured and estimated concentrations were 0.77126 and 1100.4 mg L-1, respectively. The spectra of the lettuce plants under the LED lamps were obtained at intervals of 20 min for 14 d. The time variations of the nitrate concentrations in the lettuce plants were obtained by using the regression model. The time variation could be approximated by a concavedown quadratic function. It was found that the nitrate concentration at harvest was lower in the cases in which the peak concentration occurred earlier.
Biotron Institute, 2015年, Environmental Control in Biology, 53 (4), 205 - 215, 英語[査読有り]
研究論文(学術雑誌)
研究論文(国際会議プロシーディングス)
研究論文(国際会議プロシーディングス)
[査読有り]
研究論文(学術雑誌)
Rab guanosine triphosphatases in eukaryotic cells are key regulators of membrane-trafficking events, such as exocytosis and endocytosis. Rab7 regulates traffic from early to late endosomes and from late endosomes to vacuoles/lysosomes. The Rab7-interacting lysosomal protein (RILP) was extracted from the silkworm, Bombyx mori (B. mori), and expressed in Escherichia coli (E. coli), followed by its purification. The glutathione sulfotransferase pull-down assay revealed that Rab7 of B. mori interacted with RILP of B. mori. We then produced antibodies against RILP of B. mori in rabbits for their use in Western immunoblotting and immunohistochemistry. Western immunoblotting of brain tissue for RILP revealed a single band, at approximately 50 kD. RILP-like immunohistochemical reactivity (RILP-ir) was restricted to neurons of the pars intercerebralis and dorsolateral protocerebrum. Furthermore, RILP-ir was colocalized with the eclosion hormone-ir and bombyxin-ir. However, RILP-ir was not colocalized with prothoracicotropic hormone-ir. These results were similar to those of Rab7 from our previous study. These findings suggest that RILP and Rab7 are involved in the neurosecretion in a restricted subtype of neurons in B. mori. Thus, our study is the first to report of a possible relationship between an insect Rab effector and neurosecretion.
SPRINGER, 2014年03月, HISTOCHEMISTRY AND CELL BIOLOGY, 141 (3), 311 - 320, 英語[査読有り]
研究論文(学術雑誌)
Rooftop gardening (or green roof establishment) is an urban greening afforestation method that has many environmental, economic, and urban landscaping benefits. For rooftop vegetation, stressful environments that include heat, strong winds, sunshine, and drought prevent many plants from growing well, especially in shallow soil. To establish low cost urban rooftop gardens with a low weight substrate in the summer, we evaluated the green coverage and growth of a suitable groundcover ornamental plant, blue daze (Evolvulus pilosus). Blue daze was used as rooftop vegetation in a shallow substrate (10 cm thick) on the flat rooftop of a four-story building in the middle of Kobe city, Japan, which has a temperate and humid climate. On average, the diurnal maximum temperatures and the total amounts of precipitation during each planting experiment between July and September were 30.8 degrees C and 247 mm in 2008 and 33.2 degrees C and 458 mm in 2009, respectively. Three different types of planting substrates (AS, amended soil, TM, turf mat as a dried-up blocks of sod, and FBA, furnace bottom ash) were tested to determine how they influenced plant cover, flowering, and growth. We evaluated the physical and chemical properties of the substrates that were used to establish the rooftop greeneries. In addition, the economy, ornamental and ecological values, reuse of material, and low rooftop weight burdens of the substrates were evaluated. The greatest blue daze ground cover occurred on the TM substrate (relative to the AS and FBA substrates) approximately three months after transplanting. The blue daze actively bloomed on all substrates throughout midsummer. However, alternate day irrigation largely reduced their shoot growth and flowering relative to daily irrigation. The AS and TM substrates had suitable physicochemical properties for use as planting substrates and had ideal solid, liquid, and vapor phase distributions and high water and nutrient holding capacities. FBA was a poor substrate with very low water and nutrient retention capacities, which resulted in depressed plant growth, especially under low watering conditions. To recycle FBA efficiently as a shallow low-cost planting substrate for rooftop gardening, the addition of coated fertilizers and organic matter to FBA should be considered to improve its water and nutrient holding capacities. (C) 2013 Elsevier GmbH. All rights reserved.
ELSEVIER GMBH, URBAN & FISCHER VERLAG, 2014年, URBAN FORESTRY & URBAN GREENING, 13 (1), 184 - 191, 英語[査読有り]
研究論文(学術雑誌)
Dehydration responsive element binding factors (DREBs) play important roles in plant responses to abiotic stress. In this study, a DREB2-type gene, LsDREB2A, was isolated from lettuce (Lactuca sativa L.) and characterized. LsDREB2A encoded a protein of 322 amino acid residues with the conserved apetala 2 domain and was classified into the A2 subgroup of DREBs. LsDREB2A has no sequence rich in proline, glutamic acid, serine, and threonine to trigger protein degradation. Southern blot analysis indicated that there could be more than ten DREB homologs in lettuce. A gel shift assay indicated that LsDREB2A could specifically recognize the dehydration responsive element (DRE) sequence in vitro. In a yeast one-hybrid assay, LsDREB2A bound specifically to the DRE sequence and activated the expression of both His3 and LacZ reporter genes. A transient expression experiment in lettuce protoplasts showed that LsDREB2A was localized to the nucleus. These results suggest that LsDREB2A might function as a transcription factor. Quantitative real-time polymerase chain reaction revealed that the expression of LsDREB2A was significantly increased by hyperosmotic and high salinity treatment, but not cold, heat, nor abscisic acid (ABA) treatment. Overexpression of LsDREB2A increased the tolerance of salt stress in transgenic plants. These results suggested that LsDREB2A plays an important role during high salinity stress in an ABA-independent pathway.
SPRINGER, 2014年01月, PLANT CELL TISSUE AND ORGAN CULTURE, 116 (1), 97 - 109, 英語[査読有り]
研究論文(学術雑誌)
Consumption of fresh strawberries (Fragaria xananassa) can cause oral allergy syndrome for susceptible individuals. To produce strawberries with low allergen contents a selection of hypoallergenic genotypes and the establishment of culture methods are required. Since several studies have reported that Fra a 1 is a major allergen of strawberry, the expression profiling of the Fra a 1 gene could be an indicator of the allergic potential. In this study, the expression of the Fra a 1 gene was investigated in strawberry during fruit (receptacles and achene) development. The fresh weight and anthocyan content of fruits were also measured to assess their potential correlations with Fra a 1 expression. Fruit of the strawberry were harvested for analyses from a working farm at seven different ripening stages. Real-time PCR revealed the transcript level of the Fra a 1 gene was highest at the early stage of ripening and decreased to approximately 1/70th that level by the red-colored stage. Considering that Fra a 1 belongs to the pathogenesis-related protein 10 family, this result was in line with the report that strawberry fruit may be attacked by oxygen species at the early stage of ripening. Additionally, Fra a 1 gene homologues in Fragaria vesca were surveyed to assess the inducibility of their expression. We found that a gene coding Fra a1 a2 seemed to be expressed in response to stress, and it had major stress-related elements in its promoter region. These results indicate that Fra a 1 expression can be induced by stress, and preharvest treatment to reduce the allergen content will be possible by studying the transcriptional activity of Fra a 1.
INT SOC HORTICULTURAL SCIENCE, 2014年, VII INTERNATIONAL STRAWBERRY SYMPOSIUM, 1049, 323 - 328, 英語[査読有り]
研究論文(国際会議プロシーディングス)
研究論文(国際会議プロシーディングス)
研究論文(国際会議プロシーディングス)
The effects of the exogenous application of glycinebetaine (1mM) as a drip fertigation on the root growth, fruit yield, leaf photosynthesis, stomatal conductance and chlorophyll content of glasshouse-grown tomatoes (Solanum lycopersicom L.) were compared using a crude extract (CE) and pure extract (PE) from sugarbeet in different growing seasons (ES, early summer from June to July and LS, late summer from August to September). The average air temperatures in the glasshouse were 24.5±3°C and 28±3°C in the ES and LS growing periods, respectively. The CE and PE application significantly increased and maintained the total yield of fruits, respectively, harvested in LS compared to ES. In contrast, the total yield was significantly decreased by a high incidence of blossom-end rot (BER) fruits in LS without glycinebetaine application. The CE application effectively inhibited BER and increased both the fresh weight and number of large fruits grown at a high temperature, resulting in an increase in the total yield by more than double compared to the control. The CE application also significantly enhanced root growth (both thick roots and rootlets) only in ES, whereas the PE application did not enhance root growth in either growing period. The CE-treated plants maintained photosynthesis in mature leaves at the maximum rate for approximately one month in LS, whereas the PE-treated and control plants showed significant reductions in the photosynthetic rates at the same leaf ageing. Stomatal conductance began to decrease significantly at the stage of leaf expansion at the high temperature in all of the treatments. These results indicated that CE and PE had different effects on the root growth, inhibition of BER, photosynthetic rate and stomatal conductance. CE application appears to delay leaf senescence, as based on both the significantly high values of photosynthetic O2 evolution and chlorophyll content in the leaves. By preventing BER, the CE application proved to be better than PE in improving root growth, leaf photosynthesis, chlorophyll content and yield. Thus, at low concentrations, CE can be used as an effective and low-cost organic liquid fertilizer to enhance root growth and prevent BER of fruits in glasshouse tomato production under high-temperature conditions in summer. © 2013 Elsevier B.V.
Elsevier, 2013年03月18日, Scientia Horticulturae, 152, 9 - 15, 英語[査読有り]
研究論文(学術雑誌)
[査読有り]
研究論文(国際会議プロシーディングス)
Rab proteins are small GTPases that play essential roles in vesicle transport. In this study, we examined the expression of Rab proteins and neuropeptide hormones in the brain of the silkworm, Bombyx mori. We produced antibodies against B. mori Rab1 and Rab14 in rabbits. Immunoblotting of samples of brain tissue from B. mori revealed a single band for each antibody. Rab1 and Rab14 immunohistochemical labeling in the brain of B. mori was restricted to neurons of the pars intercerebralis and dorsolateral protocerebrum. Rab1, Rab7 and Rab14 co-localized with bombyxin. Rab1 and Rab7 co-localized with eclosion hormone. Rab1 co-localized with prothoracicotropic hormone. These results suggest that Rab1, Rab7 and Rab14 may be involved in neuropeptide transport in the brain of B. mori. This is the first report on the specificity of Rab proteins for the secretion of different neuropeptides in insects.
SPRINGER, 2013年02月, HISTOCHEMISTRY AND CELL BIOLOGY, 139 (2), 299 - 308, 英語[査読有り]
研究論文(学術雑誌)
継代してから2週間から3週間培養したカルスを培地から取り出し,スペクトルの取得前に黒色ろ紙の上に置いた状態で5秒間の吸引ろ過,10秒間の吸引ろ過をしながらの洗浄処理,最後に20秒間の吸引ろ過を施すことで,培地の硝酸イオン濃度に依存せず,カルス本来の硝酸イオン濃度に近い値を測定できた.洗浄および吸引ろ過の処理を施した後のサンプルをハイパースペクトルカメラで撮影し,撮影後に硝酸イオン濃度実測値を測定した.得られた吸光スペクトルと硝酸イオン濃度から硝酸イオン濃度推定の検量線を作成した.検量線作成に用いたサンプルの硝酸イオン濃度範囲は126.0 mg L-1から2697 mg L-1であった.ビニング処理を施して取得された,526 nmから877 nmの波長の吸光スペクトルに対し,前処理としてMSCを適用し,PLS回帰分析を行って導出された検量線の硝酸イオン濃度推定精度は評価用データ相関係数が0.7363となった.
検量線作成において問題となった点は,カルスの撮影期日が同じサンプルで作成した検量線の精度は非常に高いが,異なる撮影期日のサンプルを混合すると推定精度が低下することであった.撮影期日が異なるとカルス培養の継代期日も異なるが,異なる継代期日のカルスで同様な濃度のサンプルスペクトル間に明らかなオフセットが生じていることがわかり,この現象が推定精度の低下を招いていると考えられた.そこでMSC法をスペクトルの前処理法に選択したが,推定精度の大きな向上は見られなかった.この結果から継代期日の異なるカルスは異なる性質を持った物質であると考えられた.全く同じ遺伝子を持ったカルスサンプルであるが,培養環境や撮影環境の微少な変化により細胞の状態が大きく異なっていると予想された.カルスは未分化の細胞であるため環境の変化に非常に敏感であり,温度や湿度,継代日からの経過日数,培地の状態など様々な要因の微妙な変化の影響を少しずつ受け,最終的にはカルス毎に性質が異なるサンプルとなってしまったと考えられる.
以上の考察から本研究で開発したハイパースペクトルイメージングシステムを適用する場合はカルスの培養環境や撮影環境を厳密に定め,一定に保つ必要があると考えられる.
本研究で使用したカルスは野生株であるが,組換えカルスにも本研究の成果を適用できると考える.導入予定の遺伝子はNR遺伝子のみである.薬剤耐性マーカー遺伝子などは使用しない.このため野生株の形質にNRの生産性が向上する形質が加わるだけである.従って生産量が増えるタンパク質はNRのみである.NR生産量の増減は培地の硝酸濃度や光強度などの外的環境によっても起こりえることである.NR遺伝子導入が今回作成の検量線の推定精度に影響するとは考えにくい.
[査読有り]
研究論文(学術雑誌)
To determine the sex of asparagus (Asparagus officinalis) at the seedling stage, an easy, economical, and reliable method was developed. We used a modified single-step DNA extraction protocol, which resulted in a crude extract containing sufficient genomic DNA for use as a template. The male-specific marker (Asp1-T7sp) is a dominant marker and may lead to false negatives caused by an incomplete reaction; therefore, a multiplex polymerase chain reaction (PCR) was developed using a ribosomal RNA gene marker. The resulting banding pattern distinguished males from females without false negatives. To determine the best tissue for extraction of template DNA, phylloclades (a specialized stem that resembles and functions like a leaf) or root tips of individual asparagus plants were collected and weighed. A 4.0-mg phylloclade sample or a 0.8-mg root sample provided sufficient DNA for PCR analysis of asparagus. Root excision at day 19 did not affect subsequent growth of asparagus seedlings after 28 days. The method can determine the sex of asparagus at day 19 after seeding. A combination of single-step DNA extraction from root tips and multiplex PCR made for a simple and reliable screening method.
AMER SOC HORTICULTURAL SCIENCE, 2012年02月, HORTTECHNOLOGY, 22 (1), 77 - 82, 英語[査読有り]
研究論文(学術雑誌)
Lettuce tipburn is an irreversible physiological disorder caused by calcium deficiency that decreases the crop value. Breeding a tipburn-resistant cultivar is the only causal therapy in many cases. In this study, we investigated an efficient method to evaluate lettuce resistance to tipburn in vitro. Seedlings of 19 lettuce cultivars representing three head types were cultured on agar medium containing EGTA, which chelates Ca2+. The percentage of tipburned leaves decreased proportionally with EGTA concentration. Susceptible cultivars were distinguished at 0.01 mM EGTA, whereas resistant cultivars were classified at 1.0 mM EGTA. Based on mean values of tipburn measurements, tipburn susceptibility was highest for 'Leaf Lettuce', followed by 'Butterhead Lettuce', and then 'Crisphead Lettuce'. Two cultivars were selected for further tests using hydroponic and pot culture. The rank order of susceptibility to tipburn in these experiments was consistent with that of the in vitro assay. The in vitro evaluation of lettuce susceptibility to calcium deficiency is useful for initial screening of lettuce cultivars against tipburn incidence. Resistant cultivars identified in this study are practical candidates for cultivation in controlled environments, such as a plant factory, while sensitive cultivars are also useful as indicator plants to monitor environmental conditions.
SPRINGER, 2012年02月, PLANT CELL TISSUE AND ORGAN CULTURE, 108 (2), 221 - 227, 英語[査読有り]
研究論文(学術雑誌)
Plants can synthesize some antioxidants, including L-ascorbic acid (AsA) and polyphenol, in response to environmental stresses. Antioxidants detoxify reactive oxygen species in plants and also aid in human health. In this study, we demonstrate that a novel hydroponic treatment can increase leafy vegetable nutritional quality without retarding growth. Leaf lettuce (Lactuca sativa) was grown hydroponically and subjected to rhizosphere drought stress by lowering the water level in the solution tub before harvesting. Appropriate drought stress using this method could increase AsA, polyphenol, and sugar content by 24%, 50%, and 17%, respectively, and decrease nitrate nitrogen content by 18% without reducing yield. Similar effects of drought stress on AsA content were observed in four other plant species. This hydroponic method has a universal potential to increase leafy vegetable quality without reducing yield in controlled environments such as plant factories.
AMER SOC HORTICULTURAL SCIENCE, 2012年02月, HORTTECHNOLOGY, 22 (1), 121 - 125, 英語[査読有り]
研究論文(学術雑誌)
Ii Y., URAGAMI A., UNO Y., KANECHI M., INAGAKI N., 2012. RAPD-based analysis of differences between male and female genotypes of Asparagus officinalis. Hort. Sci. (Prague), 39: 33-37. Asparagus (Asparagus officinalis L.) plants are dioecious. All-male cultivars are desired because of their higher yields. To increase the proportion of male individuals planted in the field and expedite the breeding of all-male cultivars in asparagus, development of generally applicable molecular markers to distinguish male and female individuals is required. Bulked genomic DNA samples from ten male (XY) and ten female (XX) plants was screened with 10-bp random primers. Of the 188 primers tested, the primer T35R54 produced a 1600-bp fragment observed only in male individuals. The specificity of this T35R54-1600 marker was verified using DNA from one supermale (YY) and one female (XX) breeding line and their four F-1 progenies (XY). The T35R54-1600 marker fragment was observed in both supermale and all-male lines. The sequence of the T35R54 primer (5'-TTCACGGTGG-3') was absent among the sequences of primers or amplified fragments from previous studies. Therefore, this marker could be useful as a sex-related marker in future studies to increase the reliability of sex determination in asparagus.
CZECH ACAD AGRIC SCI, 2012年, HORTICULTURAL SCIENCE, 39 (1), 33 - 37, 英語[査読有り]
研究論文(学術雑誌)
[査読有り]
研究論文(国際会議プロシーディングス)
The purpose of this research is to develop a method for non-destructive measurement of nitrate concentration in vegetable leaves. Near-infrared (NIR) spectroscopy was adopted for the measurement. Spinach and komatsuna leaves were examined. Absorption spectra at small areas in the leaves were measured by a NIR spectrometer. The measured areas were cut away from the leaves and their nitrate concentrations were measured by a liquid chromatography analyzer. Mathematical models to estimate the nitrate concentration from the spectra were developed by PCR or PLS methods with a wavelength selection algorithm. As a result, the maximum coefficient of determination between the measured and estimated concentrations became more than 0.8. © 2011 IFAC.
The INTERNATIONAL FEDERATION OF AUTOMATIC CONTROL, 2011年, IFAC Proceedings Volumes (IFAC-PapersOnline), 18 (1), 1773 - 1778, 英語[査読有り]
研究論文(国際会議プロシーディングス)
The purpose of this research is to develop a method for non-destructive measurement of nitrate concentration in vegetable leaves. Near-infrared (NIR) spectroscopy was adopted for the measurement. Spinach and komatsuna leaves were examined. Absorption spectra at small areas in the leaves were measured by a NIR spectrometer. The measured areas were cut away from the leaves and their nitrate concentrations were measured by a liquid chromatography analyzer. Mathematical models to estimate the nitrate concentration from the spectra were developed by PCR or PLS methods with a wavelength selection algorithm. As a result, the maximum coefficient of determination between the measured and estimated concentrations became more than 0.8. © 2011 IFAC.
The INTERNATIONAL FEDERATION OF AUTOMATIC CONTROL, 2011年, IFAC Proceedings Volumes (IFAC-PapersOnline), 18 (1), 1773 - 1778, 英語[査読有り]
研究論文(国際会議プロシーディングス)
Small GTPases of the Rab family are key regulators of membrane trafficking. We produced antibodies against the Rab7 protein of Bombyx mori (BRab7) in rabbits, and against the Rab11 protein of B. mori (BRab11) in mice. The antibodies recognized BRab7 and BRab11 proteins, but did not recognize other Rab proteins. Immunoblotting of samples from brain tissue of B. mori revealed a single band for each antibody. Rab11 was expressed in most tissues, whereas Rab7 was expressed in the brain, ovary, and testis. Immunohistochemical reactivity of Rab7 and Rab11 in the brain of B. mori was restricted to neurons of the pars intercerebralis and dorsolateral protocerebrum. Double-labeling experiments demonstrated that immunohistochemical reactivity of Rab7 co-localized with that of Rab11 and partially with that of Rab8. Immunohistochemical reactivity of Rab11 and Rab8 co-localized with that of PERIOD, one of the proteins associated with circadian rhythm. These findings suggest that Rab7, Rab8, and Rab11 are involved in protein transport in the neurons of the brain of B. mori and might play a role in the control of circadian rhythm. © 2010 Springer-Verlag.
2010年12月, Histochemistry and Cell Biology, 134 (6), 615 - 622, 英語[査読有り]
研究論文(学術雑誌)
[査読有り]
研究論文(国際会議プロシーディングス)
赤色光と青色光との光量子束密度比であるR/B比を0, 1, 5, 10, 20および∞(赤色光100%)の6試験区設け栽培実験を行った. 併せて室温を20℃, 22℃, 25℃の3区設定した. また22℃区においては栽培期間を1, 2, 3週間の3区設定した. 以上により栽培温度, 栽培期間により光質がレタス生体重, 乾物重および野菜中硝酸イオン濃度に与える影響を解析した.その結果乾物率が高ければ硝酸イオン濃度が低濃度となる相関関係が示されR/B比1, 5, 10が硝酸イオン濃度低減に最も適していることが明らかとなった. 室温については生体重増加量および硝酸イオン濃度で20℃区と22℃区との間に有意差はなかったが, 乾物率では20℃区の方が有意に大きくなった. このことからこの栽培システムでは20℃区が適していることが明らかになった.
日本生物環境工学会, 2010年09月, 植物環境工学, 22 (3), 140 - 147, 頁数8, 日本語[査読有り]
研究論文(学術雑誌)
alpha-Mannosidase (EC 3.2.1.24) was purified from 'Iseimo', a native variety of Dioscorea opposita Thunb. Before purification, we investigated the composition of a viscous polysaccharide that interferes with column chromatography procedures. The polysaccharide consisted mainly of mannose, and also contained uronic acid. We used the cationic detergent cetylpyridinium chloride (CPC) to remove the polysaccharide. CPC treatment decreased viscosity without affecting alpha-mannosidase activity. We purified alpha-mannosidase 2,650-fold. The optimal pH of the enzyme was 6.0 and the optimum temperature was 65A degrees C. The K (m) value for p-nitrophenyl-alpha-d-mannopyranoside was 0.35 +/- A 0.03 mM. Activity was inhibited by swainsonine but not kifunensine. The enzyme cleaved alpha-1,2 linkages preferentially to alpha-1,6 and alpha-1,3 linkages. The M (r) of purified alpha-mannosidase was estimated to be 250-260 kDa by gel filtration and native-PAGE. Four polypeptides (86, 83, 80, and 28 kDa) were detected by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. It is unclear whether the polypeptides are encoded by one gene or multiple genes. However, N-terminal sequence analysis suggested that post-translational cleavage and/or glycosylation resulted in the three large fragments, if these amino acids were encoded by the same gene. Homology searches and characterization of the enzyme's properties indicated that Iseimo alpha-mannosidase belongs to the glycoside hydrolase family 38 proteins, and to the Class II mannosidase group.
SPRINGER HEIDELBERG, 2010年07月, Acta Physiologiae Plantarum, 32 (4), 713 - 718, 英語[査読有り]
研究論文(学術雑誌)
Rab GTPases are essential for vesicular transport, whereas adenosine triphosphate (ATP) is the most important and versatile of the activated carriers in the cell. But there are little reports to clarify the connection between ATP and Rab GTPases. A cDNA clone (Rab14) from Bombyx mori was expressed in Escherichia coli as a glutathione S-transferase fusion protein and purified. The protein bound to [H-3]-GDP and [S-35]-GTPgS. Binding of [S-35]-GTP gamma S was inhibited by guanosine diphosphate (GDP), guanosine triphosphate (GTP) and ATP. Rab14 showed GTP- and ATP-hydrolysis activity. The Km value of Rab14 for ATP was lower than that for GTP. Human Rab14 also showed an ATPase activity. Furthermore, bound [H-3]-GDP was exchanged efficiently with GTP and ATP. These results suggest that Rab14 is an ATPase as well as GTPase and gives Rab14 an exciting integrative function between cell metabolic status and membrane trafficking.
ROYAL SOC, 2010年06月, BIOLOGY LETTERS, 6 (3), 379 - 381, 英語[査読有り]
研究論文(学術雑誌)
[査読有り]
研究論文(国際会議プロシーディングス)
Rooftop gardening or green roof establishment, one of the afforestation methods for urban greening, has many benefits for the environment, economy, and urban landscape. Various environmental stresses including heat, strong wind, sunshine and drought prevent most plants from growing well with extensive green roof systems. For the establishment of urban rooftop gardening in summer, we evaluated ten ornamental plant species (Evolvulus pilosus, Fragaria x ananassa, Hedera helix, Lampranthus spectabilis, Ophiopogon japonicus, Pelargonium x hortorum, Petunia x hybrida, Thymus serphyllum, Verbena x hybrida, Vinca major) to use for greening with shallow soil (10 cm depth) on the flat rooftop of a four-story building. We measured the area increase in green coverage as a total growth rate, photosynthetic ability of detached leaves, stomatal conductance of an attached leaf, soil surface and canopy air temperature. Thymus, Evolvulus, Petunia, and Fragaria are excellent plant species for rooftop gardening judging from their speedy green coverage with high growth rates (90% in Thymus, 65% in Evolvulus, 60% in both Petunia and Fragaria at maximum for about three months in summer). Thymus and Fragaria showed high performance of water saving with relatively low stomatal conductance under semi-dry conditions and did not need to be replanted every year unlike Evolvulus and Petunia. Evolvulus had the highest photosynthetic activity at 40 degrees C. Petunia and Verbena did not grow continuously but maintained active blooming without increasing their green coverage (about 60%) throughout midsummer. Lampranthus tolerated extremely hot and dry conditions on the rooftop but grew very slowly. As Vinca and Pelargonium died back due to both heat stress and strong wind, we suggested these were not suitable for rooftop gardening. All vegetations showed a 6-8 degrees C lower temperature of soil surface than that of the concrete rooftop during summer nights. We concluded that rooftop greening has a significant effect on cooling a building and saving energy for air-conditioning in summer.
JAPAN SOC HORTICULTURAL SCI, 2010年01月, JOURNAL OF THE JAPANESE SOCIETY FOR HORTICULTURAL SCIENCE, 79 (1), 69 - 76, 英語[査読有り]
研究論文(学術雑誌)
An in vitro regeneration and transient expression systems were developed for the halophyte sea aster (Aster tripolium L.), an important genetic resource for salt tolerance. Adventitious shoots were formed from both leaf explants and suspension-cultured cells in a Murashige and Skoog (MS) (Physiol Plant 15:473-497, 1962) basal salts containing 500 mg l(-1) casamino acids, and supplemented with 5.4 mu M a-naphthaleneacetic acid (NAA) and 4.7 mu M kinetin to the culture medium. Hyperhydricity of shoots was avoided by increasing the ventilation of the culture vessel. Root formation from shoots was promoted in the presence of 26.9 mu M NAA. A high yield of protoplasts was isolated using 1% cellulase and 0.25% pectinase from both leaf mesophyll and suspension-cultured cells, and these were used for transient expression. The highest level of transient expression of the green fluorescent protein was obtained with 1 x 10(5) protoplasts ml(-1), 25 mu g batch(-1) of plasmid vector, and 30% polyethylene glycol 4,000.
SPRINGER, 2009年09月, PLANT CELL TISSUE AND ORGAN CULTURE, 98 (3), 303 - 309, 英語[査読有り]
研究論文(学術雑誌)
Calcium-dependent protein kinases (CDPKs) are essential sensor-transducers of calcium signaling pathways in plants. Functional characterization of CDPKs is of great interest because they play important roles during growth, development, and in response to a wide range of environmental stimuli. The Arabidopsis genome encodes 34 CDPKs, but very few substrates of these enzymes have been identified. In this study, we exploited the unique characteristics of CDPKs to develop an efficient approach for the discovery of CDPK-interacting proteins. High-throughput, semi-automated yeast two-hybrid interaction screens with two different cDNA libraries each containing 18 million prey clones were performed using catalytically impaired and constitutively active AtCPK4 and AtCPK11 variants as baits. The use of the constitutively active versions of the CPK baits improved the recovery of positive interacting proteins relative to the wild type kinase. Titration of interaction strength by growth under increasing concentrations of 3-aminotriazole (3-AT), a histidine analog and competitive inhibitor of the His3 gene product, confirmed these results. Possible mechanisms for this observed improvement are discussed. The reproducibility of this approach was assessed by the overlap of several interacting proteins of AtCPK4 and AtCPK11 and the recovery of several putative substrates and indicated that yeast two-hybrid screens using constitutively active and/or catalytically impaired forms of CDPK provides a useful tool to identify potential substrates of the CDPK family and potentially the entire protein kinase superfamily.
SPRINGER HEIDELBERG, 2009年04月, MOLECULAR GENETICS AND GENOMICS, 281 (4), 375 - 390, 英語[査読有り]
研究論文(学術雑誌)
[査読有り]
研究論文(学術雑誌)
Small GTPases of the Rab family act as essential regulators of vesicle transport pathways. Five Rab cDNA clones (BRab1, 7, 8, 11 and 14) from Bombyx mori were expressed in Escherichia coli as a thioredxin or glutathione sulfotransferase fusion protein. After purification, the fusion protein was tested for phosphorylation using protein kinase C (PKC). Results indicate that all of them were phosphorylated in vitro. The phosphorylation site of BRab1 was determined by mass-spectrometric analysis, which identified that Ser-17 of BRab1 was phosphorylated by PKC. Deletion and site-directed mutagenesis indicated that Ser-111 of BRab8, in addition to Ser-17, was newly phosphorylated. Further immunohistochemical analysis using antibodies against Rab8 indicated that there are some Rab8 immunoreactive cells close to the neuropeptide secreting cells. This result suggests that in insects Rab proteins are regulated by phosphorylation and at least some of them are involved in neuropeptide secretion.
CZECH ACAD SCI, INST ENTOMOLOGY, 2009年, EUROPEAN JOURNAL OF ENTOMOLOGY, 106 (4), 499 - 506, 英語[査読有り]
研究論文(学術雑誌)
Ploidy levels of in vitro shoots produced through anther culture of asparagus were determined by flow cytometry (FCM) analysis instead of counting the chromosome number of root tips that were difficult to regenerate. Of the I 10 anther-derived shoots, there were 83 diploids, 21 tetraploids, 3 octaploids, and 3 mixoploids, but no haploids. When flower buds of different length were used as starting material for anther culture, the richest variation in polyploidy originated from short flower buds (1.0-1.5 mm). This result suggested that polyploids might develop by the fusion of nuclei during pollen-mother cell division before the tetrad stage. We used scanning electron microscopy (SEM) to measure the stomatal length of in vitro and field-grown shoots in order to clarify the differences among polyploids determined by FCM analysis. When the developmental stage of tissue was taken into account, we found statistically significant differences in stomatal length among all polyploids (diploid, tetraploid, and octaploid) for both stems and phyllodes (P<0.001). Stomata on stems were longer than those on phyllodes, and mature tissue had longer stomata than immature tissue. Morphological observations with SEM found that immature tissue had guard cells set below epidermal cells, and were poorly developed. Field-grown diploids had much shorter stornatal length than in vitro diploids. Male plants grown in the field had significantly longer stomata than female plants in both their immature stems and mature phyllodes (P<0.001). These results indicated that stomatal length can be a reliable ploidy index by removing any other influential factors such as sex, environment and developmental stage.
JAPAN SOC HORTICULTURAL SCI, 2009年01月, JOURNAL OF THE JAPANESE SOCIETY FOR HORTICULTURAL SCIENCE, 78 (1), 103 - 108, 英語[査読有り]
研究論文(学術雑誌)
We established a novel in vitro method, termed the root recovery assay, to evaluate the survival under osmotic stress of lettuce (Lactuca sativa L.) seedlings. Under salinity and drought stress, combination of the root-bending assay and root recovery assay showed the same trends in dry weight and survival rate as a hydroponic culture. Both in vitro assays and hydroponics ranked the three lettuce cultivars in the same order of drought tolerance. The root-bending assay evaluated the plant's growth and the root recovery assay indicated the plant's survival. In addition, the combined assay required less space and approximately half the time period compared with the hydroponic culture. These results suggested that application of the root-bending and root recovery assay should be a rapid and space-saving method with which to evaluate the osmotic stress tolerance of lettuce from both growth and survival standpoints.
SPRINGER, 2008年10月, PLANT CELL TISSUE AND ORGAN CULTURE, 95 (1), 101 - 106, 英語[査読有り]
研究論文(学術雑誌)
P450 (cytochrome P450) enzymes catalyse the mono-oxygenation of a wide range of compounds such as steroids, fatty acids, vitamins and drugs. In the present paper we demonstrate a system for bioconverting diverse compounds [flavanone, DHEA (dehydroepiandrosterone) and 7-ethoxycoumarin] using P450 species expressed in Escherichia coli. First, we expressed four P450 species: rabbit CYP2B (MO family 2, subfamily B), fruitfly (Drosophila) CYP317A, rat CYP3A23 and mouse CYP2J5. Next, we added substrates directly to the incubation medium. The resulting metabolites were extracted and analysed by HPLC and spectrofluorimetry. The first substrate, 7-ethoxycoumarin, was de-ethylated by CYP2B; CYP2J5 and CYP3A23 showed weak activity, and CYP317A had no activity for 7-ethoxycoumarin. We next used flavanone, a flavonoid, as a substrate for these four MO species and other P450 species expressed previously. As a result, CYP2B, CYP2C43 and CYP2C29 catalysed flavanone 2-hydroxylation. CYP2A5 catalysed 2- and 4-hydroxylations. Finally, to produce diverse modified compounds, variants of CYP2A5 with point mutations were incubated with a steroid (DHEA) and an antioxidant (flavanone) in vivo. HPLC analysis indicated that two P450 species produced a 7-beta-hydroxy-DHEA and two P450 species produced a 2-alpha-hydroxy-DHEA. Four P450 species catalysed flavanone 2- and 4-hydroxylations. These results indicate that bioconversion by P450 is a useful technique to modify small molecules (steroids, coumarin and flavanone) and produce new, diverse hydroxylated compounds, which could be used for high-throughput screening for drug discovery.
WILEY, 2008年08月, BIOTECHNOLOGY AND APPLIED BIOCHEMISTRY, 50, 165 - 171, 英語[査読有り]
研究論文(学術雑誌)
Small GTPases of the Rab family are key regulators of membrane trafficking. Monoclonal antibodies are useful tools for identifying proteins that interact with other proteins and for examining their tissue distribution. We selected a monoclonal antibody against Rab8 of Bombyx Mori L. It specifically recognized amino acid residues 30-109, which are conserved among Rab8 proteins, and did not recognize any other Rab proteins. Western blotting using the antibody revealed one band in the brains of B. Mori and rat. Far-Western blotting analysis detected three proteins interacting with Rab8. These results indicate that this antibody is useful for clarifying the physiological function of Rab8 of B. Mori and other species. This is a report of a study on a monoclonal antibody against insect Rab protein.
CZECH ACAD SCI, INST ENTOMOLOGY, 2007年10月, EUROPEAN JOURNAL OF ENTOMOLOGY, 104 (4), 641 - 645, 英語[査読有り]
研究論文(学術雑誌)
Rooftop gardening, one of the methods for urban greening, has many benefits for the environment, economy, and urban landscape. The research for green roof establishment is most prevalent in Germany, but is still under development in Japan. Extensive green roof systems with light and shallow substrates enable plants to grow without significant changes in building structure, but suitable plant species must be selected. Various environmental stresses prevent most plants from growing well in extensive green roof systems. For the establishment of urban rooftop gardening in Japan, information on what kind of plant is suitable for growing on the rooftop where environmental conditions are rather severe to many plant species must be obtained. Today, some species of turf and sedum, which are quite tolerant to dry and hot conditions like rooftops, are generally used as plant species for rooftop gardening in Japan, but they are not attractive for urban landscape. The objective of this study was to identify beautiful ornamental plants which survive and grow well on the rooftop, especially in the summer season in Japan. Five plant species (turf, thyme, sedum, verbena, and marigold) were established on the flat rooftop of the four-storey Agricultural Department building of Kobe University in summer 2005. Each plant species was grown in two kinds of shallow substrate containing perlite or not with intensive or extensive watering (5 L every other day or a week). To evaluate the suitability for rooftop gardening, the rate of plant growth, the ratio of ground cover with plant, the number of flowers and the ability of photosynthesis were measured. With limited watering, only sedum grew well. The other species grew with intensive watering. Marigolds were blown away by strong wind. Verbena and thyme appear to be suitable planting for rooftop gardening because of their eye filling and harvesting benefits. Further, thyme needs little management in comparison with verbena.
INT SOC HORTICULTURAL SCIENCE, 2007年, PROCEEDINGS OF THE INTERNATIONAL SYMPOSIUM ON HORTICULTURAL PLANTS IN URBAN AND PERI-URBAN LIFE, 762 (762), 333 - +, 英語[査読有り]
研究論文(国際会議プロシーディングス)
Auxin transport plays a significant role modifying plant growth and development in response to environmental signals such as light and gravity. However, the effect of humidity on auxin transport is rarely documented. It is shown here that the transport of labelled indole-3-acetic acid (IAA) from the shoot to the root is accelerated in rice (Oryza sativa L. ssp. indica cv. IR8) seedlings grown under saturated humidity (SH-seedlings) compared with plants grown under normal humidity (NH-seedlings). The development of lateral roots in SH-seedlings was greatly enhanced compared with NH-seedlings. Removal of the shoot from SH-seedlings reduced the density of lateral roots, and the application of IAA to the cut stem restored the lateral root density, while the decapitation of NH-seedlings did not alter lateral root development. Phloem-based auxin transport appeared responsible for enhanced lateral root formation in SH-seedlings since (i) the rate of IAA transport from the shoot to the root tip was greater than 3.5 cm h(-1) and (ii) naphthylphthalamic acid (NPA)-induced reduction of polar auxin transport in the shoot did not influence the number of lateral roots in SH-seedlings. It is proposed that high humidity conditions accelerate the phloem-based transport of IAA from the leaf to the root, resulting in an increase in the number of lateral roots.
OXFORD UNIV PRESS, 2007年, JOURNAL OF EXPERIMENTAL BOTANY, 58 (7), 1695 - 1704, 英語[査読有り]
研究論文(学術雑誌)
Anthocyanins are responsible for many of the vivid colors in higher plants. The natural floral color in the petals of Rosa hybrida 'Charleston' undergoes a striking change from yellow to red over a period of 10-12 days. This color change is due to the accumulation of two anthocyanins, cyanidin 3-glucoside (chrysanthemin) and cyanidin 3,5-diglucoside (cyanin). The production of anthocyanins is determined by expression of at least four genes, namely dihydroflavonol 4-reductase (DFR), anthocyanidin synthase (ANS), flavonoid 3-0-glucosyltransferase (MG7), and flavonoid 5-O-glucosyltransferase (UF5GT). The cDNA fragments of these genes were cloned from rose and the deduced amino acid sequences of these genes shared 51 similar to 97% identities with the corresponding sequences from other plant species. Rose flowers were exposed to different light conditions such as white light (FL, cool-white fluorescent lamp) or ultraviolet radiation (UVB, 290-320 nm; UVA, 320-380 nm) to promote the accumulation of the anthocyanins in petals during flower unfolding. Anthocyanin contents were measured by HPLC and the gene expression was investigated by RNA gel-blot analysis and RT-PCR. The expression of these genes increased dramatically as flowers unfolded and also correlated positively with the production of anthocyanins. The expression of DFR was detectable in early stages of flower unfolding and the highest expression of ANS, UF3GT and UF5GT was detected during the latter half of flower unfolding. The expression levels of these genes and the temporal buildup of anthocyanins in petals were stimulated by exposing them to UVB or FL+UVB radiation. These results show the relationship between the expression of anthocyanin biosynthetic genes and the light-signaling stimuli within the narrow range of UV wavelengths. UVB radiation is increasing gradually due to the rapid destruction of ozone layer.
INTERNATIONAL SOCIETY HORTICULTURAL SCIENCE, 2007年, Acta Horticulturae, 760 (760), 643 - 650, 英語[査読有り]
研究論文(国際会議プロシーディングス)
Calcium-dependent protein kinases (CDPKs) are a novel class of signaling molecules that have been broadly implicated in relaying specific calcium-mediated responses to biotic and abiotic stress as well as developmental cues in both plants and protists. Calcium-dependent autophosphorylation has been observed in almost all CDPKs examined, but a physiological role for autophosphorylation has not been demonstrated. To date, only a handful of autophosphorylation sites have been mapped to specific residues within CDPK amino acid sequences. In an attempt to gain further insight into this phenomenon, we have mapped autophosphorylation sites and compared these phosphorylation patterns among multiple CDPK isoforms. From eight CDPKs and two CDPK-related kinases from Arabidopsis thaliana and Plasmodium falciparum, 31 new autophosphorylation sites were characterized, which in addition to the previously described sites, allowed the identification of five conserved loci. Of the 35 total sites analyzed approximately one-half were observed in the N-terminal variable domain. Homology models were generated for the protein kinase and calmodulin-like domains, each containing two of the five conserved sites, to allow intelligent speculation regarding subsequent lines of investigation.
WILEY-V C H VERLAG GMBH, 2006年06月, Proteomics, 6 (12), 3649 - 3664, 英語[査読有り]
研究論文(学術雑誌)
Calcium-dependent protein kinases (CDPKs) are sensor-transducer proteins capable of decoding calcium signals in diverse phosphorylation-dependent calcium signaling networks in plants and some protists. Using a novel yeast two-hybrid (YTH) approach with constitutively active and/or catalytically inactive forms of AtCPK11 as bait, we identified AtDi19 as an AtCPK11-interacting protein. AtDi19 is a member of a small family of stress-induced genes. The interaction was confirmed using pull-down assays with in vitro translated AtCPK11 and GST-AtDi19 and localization studies in Arabidopsis protoplasts cotransfected with AtCPK11:GFP and AtDi19:DsRed2 protein fusions. We further showed that the interaction of AtDi19 is specific to both AtCPK4 and AtCPK11, whereas other closely related CPKs from Arabidopsis interacted weakly (e.g., AtCPK12) or did not interact (e.g., AtCPK26, AtCPK5 and AtCPK1) with AtDi19. Deletion analyses showed that a region containing two predicted nuclear localization signals (NLS) and a nuclear export signal (NES) of AtDi19 is essential for interaction with AtCPK11. We further demonstrated that AtDi19 is phosphorylated by AtCPK11 in a Ca2+-dependent manner at Thr105 and Ser107 within the AtDi19 bipartite NLS using in vitro kinase assays. Our data suggest that disruption of the autoinhibitor domain leading to the formation of a constitutively active CDPK may stabilize kinase-substrate inter-actions without affecting specificity. (c) 2006 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.
ELSEVIER SCIENCE BV, 2006年02月, FEBS LETTERS, 580 (3), 904 - 911, 英語[査読有り]
研究論文(学術雑誌)
The ultraviolet-B radiation (UV-B: 280 to 320 nm) promotes the accumulation of the anthocyanins in rose plants. Open flowers of Rosa hybrida cultivars 'Charleston' and 'Ehigasa' undergo a striking color change from yellow to red over 10-12 days under natural daylight due to the accumulation of the anthocyanins in the petals. The anthocyanin present in the rose petals was identified as cyanidin 3-glucoside (chrysanthemin) and cyanidin 3,5-diglucoside (cyanin) by using paper chromatography, partial acid hydrolysis, and HPLC. To determine the role of UV-B in the anthocyanin biosynthesis pathway, rose plants were grown under different light conditions supplemented with or without UV-B. Anthocyanin accumulation was suppressed in the treatment without UV-B and the accumulation started following to exposure to UV-B by a gradual increase of cyanin, followed by a dramatically increase of chrysanthemin. Furthermore, it revealed that the unfolding flower stage 3 to 4 (about 50% anthesis) was critical for anthocyanin biosynthesis in rose.
Biotron Institute, 2006年, Enviornment Control in Biology, 44 (2), 103 - 110, 英語[査読有り]
研究論文(学術雑誌)
A suspension cultured cell line was established from the cultivar of Rosa hybrida 'Charleston' as a study model to understand the response of the anthocyanin biosynthesis pathway to environmental cues. The major identified anthocyanin in cell cultures was cyanidin 3-glucoside (chrysanthemin). The anthocyanin yield was enhanced by culturing cells in the EM medium with added sucrose at high concentration under additional UV-B radiation to white light. Three cDNA fragments were cloned with degenerate primers by RT-PCR and the obtained sequences shared high homology with putative key enzymes (DFR, ANS, and UF3GT) of other species. The expression levels of these three genes were promoted under optimum conditions for anthocyanin accumulation. These results suggest that expression levels of these genes were closely correlated with a temporal buildup of anthocyanins in response to environmental factors. Copyright © 2006 The Japanese Society for Plant Cell and Molecular Biology.
Japanese Society for Plant Cell and Molecular Biology, 2006年, Plant Biotechnology, 23 (4), 379 - 385, 英語[査読有り]
研究論文(学術雑誌)
To isolate and analyze salt-stress inducible genes in a halophyte, sea aster (Aster tripolium L.), we screened 5760 Arabidopsis cDNA clones by macroarray procedure using 33P-labeled cDNA targets synthesized from mRNAs isolated from NaCl treated and untreated sea aster seedlings. Seventeen Arabidopsis cDNAs were hetero - hybridized to NaCl inducible sea aster genes. These cDNAs were used as probes to isolate cDNA homologs from a sea aster cDNA library. One of the obtained cDNAs shared 71% amino acids identity with Arabidopsis cysteine protease (AtCysP) and named SaCysP (sea aster CysP). Northern blot analysis revealed that mRNAs corresponding to both SaCysP and AtCysP were induced by salt and osmotic stress in leaves. On the other hand, SASR21 mRNA encoding another CysP in sea aster was irresponsive to these stress in leaves but respond in roots. SaCysP and SASR21 genes may have a tissue-specific function in stress response by modulating their expression levels.
Japanese Society for Plant Cell and Molecular Biology, 2004年, Plant Biotechnology, 21 (2), 127 - 133, 英語[査読有り]
研究論文(学術雑誌)
To understand the mechanisms of salt tolerance in a halophyte, sea aster (Aster tripolium L.), we studied the changes of water relation and the factors of photosynthetic limitation under water stress and 300 mM NaCl stress. The contents of Na+ and Cl- were highest in NaCl-stressed leaves. Leaf osmotic potentials (Psi(s)) were decreased by both stress treatments, whereas leaf turgor pressure (Psi(t)) was maintained under NaCl stress. Decrease in Psi(s) without any loss of Psi(t) accounted for osmotic adjustment using Na+ and Cl- accumulated under NaCl stress. Stress treatments affected photosynthesis, and stomatal limitation was higher under water stress than under NaCl stress. Additionally, maximum CO2 fixation rate and O-2 evolution rate decreased only under water stress, indicating irreversible damage to photosynthetic systems, mainly by dehydration. Water stress severely affected the water relation and photosynthetic capacity. On the other hand, turgid leaves under NaCl stress have dehydration tolerance due to maintenance of Psi(t) and photosynthetic activity. These results show that sea aster might not suffer from tissue dehydration in highly salinized environments. We conclude that the adaptation of sea aster to salinity may be accomplished by osmotic adjustment using accumulated Na+ and Cl-, and that this plant has typical halophyte characteristics, but not drought tolerance.
SPRINGER-VERLAG TOKYO, 2003年02月, JOURNAL OF PLANT RESEARCH, 116 (1), 65 - 70, 英語[査読有り]
研究論文(学術雑誌)
In order to understand responses of the halophyte sea aster (Aster tripolium L.) to salt stress at the molecular level, we used a differential screening strategy. Nine cDNAs of genes induced by salt stress were cloned by this method from sea aster. The clones were collectively named SASR (sea aster stress responsive). These cDNAs were mainly associated with osmoprotectant synthesis, protein metabolism, protein degradation or signal transduction. All the mRNAs corresponding to the SASR genes were accumulated under high salt conditions and the transcripts of 3 out of 9 genes were induced by osmotic stress.
Japanese Society for Plant Cell and Molecular Biology, 2003年, Plant Biotechnology., 20(4),pp.317-322 (4), 317 - 322, 英語[査読有り]
研究論文(学術雑誌)
Asparagus are dioecious and perennial, being grown for 10 years or more. All male cultivars are desired due to higher yields and homogenity, and vigor than cultivars. We attempted to obtain polyembryonic seeds in which haploid seedling might be caught. In addition, searching peculiar genetic markers was attempted on a female and a male bulk and haploid-derived calluses by RAPD (Random Amplified Polymorphic DNA) analysis. The frequency of polyembryonic seeds was 0.34%(34/9925). All polyembryonic seeds had twin seedlings except which one embryonic seed had triplet seedlings. The frequency of polyembryonic seeds was suggested to be different according to the genotypes. The survived 33 seedlings were composed of one haploid (2n=x=10), one triploid (2n=3x=30), and 31 diploids (2n=2x=20). Cladophylls had larger stomata in order of haploid, diploid, and triploid. The size of stomata was less different between haploid and diploid, while larger difference was observed between haploid or diploid, and triploid. RAPD analysis with ten kinds of random primers resulted in occurrence of 3 bands specific to the male bulk and the calluses with the primer No. 1, 3 and 5, and one band specific to the female bulk with primer No. 5.
INTERNATIONAL SOCIETY HORTICULTURAL SCIENCE, 2002年, PROCEEDINGS OF THE TENTH INTERNATIONAL ASPARAGUS SYMPOSIUM, 589 (589), 217 - 224, 英語[査読有り]
研究論文(国際会議プロシーディングス)
The induction of the dehydration-responsive Arabidopsis gene, rd29B, is mediated mainly by abscisic acid (ABA), Promoter analysis of rd29B indicated that two ABA-responsive elements (ABREs) are required for the dehydration-responsive expression of rd29B as cis-acting elements. Three cDNAs encoding basic leucine zipper (type)type ABRE-binding proteins were isolated by using the yeast one-hybrid system and were designated AREB1, AREB2, and AREB3 (ABA-responsive element binding protein). Transcription of the AREB1 and AREB2 genes is up-regulated by drought, NaCl, and ABA treatment in vegetative tissues. In a transient transactivation experiment using Arabidopsis leaf protoplasts, both the AREB1 and AREB2 proteins activated transcription of a reporter gene driven by ABRE, AREB1 and AREB2 required ABA for their activation, because their transactivation activities were repressed in aba2 and abi1 mutants and enhanced in an era1 mutant. Activation of AREBs by ABA was suppressed by protein kinase inhibitors. These results suggest that both AREB1 and AREB2 function as transcriptional activators in the ABA-inducible expression of rd29B, and further that ABA-dependent posttranscriptional activation of AREB1 and AREB2, probably by phosphorylation. is necessary for their maximum activation by ABA, Using cultured Arabidopsis cells, we demonstrated that a specific ABA-activated protein kinase of 42-kDa phosphorylated conserved N-terminal regions in the AREB proteins.
NATL ACAD SCIENCES, 2000年10月, PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 97 (21), 11632 - 11637, 英語[査読有り]
研究論文(学術雑誌)
Six partial PCR fragments that encode aquaporin homologues in the halophyte sea aster Aster tripolium were cloned using genomic DNA and cDNA as templates. The deduced amino acid sequences were highly similar to those of other major intrinsic proteins (MIPs) in plants. The expression patterns of sea aster mip genes in leaves and suspension-cultured cells during a 24-hr NaCl stress period were studied with PCR samip (sea aster major intrinsic protein) fragments. Three samip (A-C) fragments were obtained by PCR with genomic DNA as templates, and 3 samip (D-F) fragments were obtained by RT-PCR Among the 6 genes, expression of Samip A, B and C was significantly inducible by salt stress. The level of Samip A, B and C mRNAs was dependent on NaCl concentration in both whole plants and cultured cells. However, no significant accumulation of Samip transcripts was observed in salt-adapted cells. These observations suggest that Samip A, B and C genes play an important role in the early salt-adaptable stage in sea asters.
BOTANICAL SOC JAPAN, 1998年09月, JOURNAL OF PLANT RESEARCH, 111 (1103), 411 - 419, 英語[査読有り]
研究論文(学術雑誌)
野菜類の中でも高耐塩性をもつとされるアスパラガス(Asparagus officinalis L.)およびテーブルビート(Beta vulgaris L.)と,汽水域に生息し,塩ストレスに対して有効な遺伝資源をもつ塩生植物ウラギク(Asterti tipolium L.)を用いて,0~300mMの様々の塩ストレス下で発芽の状況を調査するとともに,水耕栽培を行い,栄養生長期における生長量や形態的な差異を調査した結果,以下のような知見を得た.
アスパラガスおよびテーブルビートは,0 mM NaClの対照区で90%近くが発芽したが,50mMのNaCI条件下での発芽率は,それぞれ,50%および61%,また100mMでは,12%および21%であった.ウラギクについても,塩濃度が増すにつれて発芽は抑制されたが,150mMのNaCl処理でも,対照区の発芽率を100とした時の50%以上が発芽しており,種子段階からの高耐塩性が示唆された.
アスパラガスおよびテーブルビート実生を目的の塩濃度(50および100mM)で直接処理した場合,植物体は萎ちょうおよび枯死したため,この二つの植物については段階的に濃度を上げて処理した.塩ストレスを与えた栄養生長期の植物では,草丈には差がなかったが,塩処理区の根長が対照区に較べて有意に長くなった点は供試した植物に共通した.アスパラガスおよびテーブルビートの地下部の乾物重は100mMのNaClによって抑制された.一方,ウラギクは,目的の塩濃度で直接処理したにもかかわらず,乾物生産は,150mMのNaClでは対照区とほぼ同様で,300mMのNaClで抑制された.
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研究論文(学術雑誌)
We established salt tolerant cell lines, which survived and grew under high salinity conditions with 150 mM (S-150) and 300 mM (S-300) NaCl, to study the effects of salt stress on the proliferation and protein profile of these cells in the halophyte sea aster, Aster tripolium L. These salt-adapted cell lines were produced from leaves and selected by repeated suspension subculture in media containing NaCl every 25 days for five cycles. S-150 cells displayed no inhibition in their growth compared to control cells maintained under non-stressed conditions. S-150 cells exhibited approximately a 15-fold increase in both fresh and dry weight during the 25 days under saline conditions. S-300 cells showed positive growth under severe salt stress, but their dry matter gain was significantly less than that of the S-150 cells, with only a 2.5-fold increase in dry weight. We also detected changes in the protein profile of salt-adapted cells with two specifically induced polypeptides (basic 58.4 and acidic 24.8 kDa) and one enhanced polypeptide (basic 15.1 kDa) in the soluble fraction, and one specifically induced polypeptide (42.0 kDa) in the insoluble fraction.
BOTANICAL SOC JAPAN, 1996年12月, JOURNAL OF PLANT RESEARCH, 109 (1096), 409 - 414, 英語[査読有り]
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Plants respond and adapt to environmental changes including drought, high salinity and low temperature. Abscisic acid (ABA) plays important roles in these stress responses. A number of plant genes are induced by water stress, such as drought, high salinity and low temperature, and are thought to function in the stress tolerance and responses of the plant. At least four signal transduction pathways control these genes in Arabidopsis thaliana: two are ABA-dependent and two are ABA-independent A cis-acting element named DRE (Dehydration Responsive Element) is involved in one of the ABA-independent signal transduction pathways, and its DNA binding proteins have been characterized. Drought- and ABA-inducible MYC and MYB homologues are involved in ABA-responsive gene expression in Arabidopsis. Roles of these cis and transacting factors in water stress responses are discussed. In addition, a number of genes for protein kinases, enzymes involved in phosphatidyl inositol metabolism (PI turnover) and transcription factors are also induced by water stress. and thought to be involved in the stress signal transduction cascades. Possible signaling processes in water stress response are discussed.
BOTANICAL SOC JAPAN, 1998年06月, JOURNAL OF PLANT RESEARCH, 111 (1102), 345 - 351, 英語[査読有り]
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