Wiley Acta Medica Okayama 1936-5209 19 3 2025 Oregon Wolfe barley genetic stocks – Research and teaching tools for next generation scientists e70004 EN Margaret R. Krause Department of Crop and Soil Science, Oregon State University Juan David Arbelaez Department of Crop Sciences, University of Illinois at Urbana-Champaign Åsmund Asdal Nordic Genetic Resource Centre Ramzi Belkodja CIHEAM-Zaragoza Nancy Boury Department of Plant Pathology, Entomology, and Microbiology, Iowa State University Victoria C. Blake Department of Plant Sciences and Plant Pathology, Montana State University Patrick J. Brown Department of Plant Sciences, University of California-Davis Ana Casas Departamento de Genética y Producción Vegetal, Estación Experimental Aula Dei–CSIC Luis Cistué Departamento de Genética y Producción Vegetal, Estación Experimental Aula Dei–CSIC Alba Farré�]Martínez AGROTECNIO-CERCA Center, Universidad de Lleida Scott Fisk Department of Crop and Soil Science, Oregon State University Gregory S. Fuerst U.S. Department of Agriculture-Agricultural Research Service, Corn Insects and Crop Genetics Research Unit, Iowa State University Estela Giménez Department of Biotechnology-Plant Biology, School of Agricultural, Food and Biosystems Engineering, Universidad Politécnica de Madrid Carla Guijarro�]Real Department of Biotechnology-Plant Biology, School of Agricultural, Food and Biosystems Engineering, Universidad Politécnica de Madrid Katy Guthrie Department of Agronomy and Plant Genetics, University of Minnesota Margaret Halstead Aardevo North America Laura Helgerson Department of Crop and Soil Science, Oregon State University Hiroshi Hisano Institute of Plant Science and Resources, Okayama University Ernesto Igartua Departamento de Genética y Producción Vegetal, Estación Experimental Aula Dei–CSIC Morten Lillemo Department of Plant Sciences, Norwegian University of Life Sciences Marina Martínez�]García Department of Biotechnology-Plant Biology, School of Agricultural, Food and Biosystems Engineering, Universidad Politécnica de Madrid Mariona Martínez�]Subirà AGROTECNIO-CERCA Center, Universidad de Lleida Susan McCouch Plant Breeding and Genetics Section, School of Integrative Plant Science, Cornell University Laurie McGhee Colfax-Mingo Community High School Travis Nickols Department of Crop and Soil Science, Oregon State University Nick Peters Department of Plant Pathology, Entomology, and Microbiology, Iowa State University Raymond Porter Haupert Institute for Agricultural Studies, Huntington University Ignacio Romagosa AGROTECNIO-CERCA Center, Universidad de Lleida Anja Karine Ruud Department of Plant Sciences, Norwegian University of Life Sciences Kazuhiro Sato Institute of Plant Science and Resources, Okayama University Silvio Salvi Department of Agricultural and Food Sciences, University of Bologna Giuseppe Sangiorgi Department of Agricultural and Food Sciences, University of Bologna Rebekka Schüller Department of Crop Sciences, University of Illinois at Urbana-Champaign Taner Z. Sen Crop Improvement and Genetics Research Unit, U.S. Department of Agriculture-Agricultural Research Service José Miguel Soriano AGROTECNIO-CERCA Center, Universidad de Lleida Robert M. Stupar Department of Agronomy and Plant Genetics, University of Minnesota To�]Chia Ting Agronomy Department, Purdue University Kelly Vining Department of Crop and Soil Science, Oregon State University Maria von Korff Institute of Plant Genetics, Heinrich-Heine-Universität Düsseldorf Agatha Walla Institute of Plant Genetics, Heinrich-Heine-Universität Düsseldorf Diane R. Wang Agronomy Department, Purdue University Robbie Waugh Division of Plant Sciences, School of Life Sciences, University of Dundee Roger P. Wise Department of Plant Pathology, Entomology, and Microbiology, Iowa State University Robert Wolfe Agriculture and Agri-Food Canada Eric Yao Crop Improvement and Genetics Research Unit, U.S. Department of Agriculture-Agricultural Research Service Patrick M. Hayes Department of Crop and Soil Science, Oregon State University The Oregon Wolfe Barley (OWB) mapping population (Reg. no. MP-4, NSL 554937 MAP) is a resource for genetics research and instruction. The OWBs are a set of doubled haploid barley (Hordeum vulgare L.) lines developed at Oregon State University from the F1 of a cross between Dr. Robert Wolfe's dominant and recessive marker stocks. Exhibiting a high level of genetic and phenotypic diversity, the OWBs are used throughout the world as a research tool for barley genetics. To date, these endeavors have led to 56 peer-reviewed publications, as well as three reports in the Barley Genetics Newsletter. At the same time, the OWBs are widely used as an instructor resource at the K–12, undergraduate, graduate, and professional levels. They are currently used at universities and/or institutes in German, Italy, Norway, Spain, and the United States and are currently being developed further for educational use in other countries. Genotype and phenotype data, lesson plans, and seed availability information are available herein and online. No potential conflict of interest relevant to this article was reported.

MDPI Acta Medica Okayama 2075-1729 15 2 2025 Distinct Infection Mechanisms of Rhizoctonia solani AG-1 IA and AG-4 HG-I+II in Brachypodium distachyon and Barley 235 EN Niranjan Mahadevan Graduate School of Environmental, Life, Natural Science and Technology, Okayama University Rozi Fernanda Graduate School of Environmental, Life, Natural Science and Technology, Okayama University Yusuke Kouzai Crop Stress Management Group, Division of Plant Molecular Regulation Research, Institute of Agrobiological Sciences, National Agriculture and Food Research Organization (NARO) Natsuka Kohno Faculty of Agriculture, Okayama University Reiko Nagao Faculty of Agriculture, Okayama University Khin Thida Nyein Graduate School of Environmental, Life, Natural Science and Technology, Okayama University Megumi Watanabe Graduate School of Environmental, Life, Natural Science and Technology, Okayama University Nanami Sakata Graduate School of Environmental, Life, Natural Science and Technology, Okayama University Hidenori Matsui Graduate School of Environmental, Life, Natural Science and Technology, Okayama University Kazuhiro Toyoda Graduate School of Environmental, Life, Natural Science and Technology, Okayama University Yuki Ichinose Graduate School of Environmental, Life, Natural Science and Technology, Okayama University Keiichi Mochida RIKEN Center for Sustainable Resource Science Hiroshi Hisano Institute of Plant Science and Resources, Okayama University Yoshiteru Noutoshi Graduate School of Environmental, Life, Natural Science and Technology, Okayama University Rhizoctonia solani is a basidiomycete phytopathogenic fungus that causes rapid necrosis in a wide range of crop species, leading to substantial agricultural losses worldwide. The species complex is divided into 13 anastomosis groups (AGs) based on hyphal fusion compatibility and further subdivided by culture morphology. While R. solani classifications were shown to be independent of host specificity, it remains unclear whether different R. solani isolates share similar virulence mechanisms. Here, we investigated the infectivity of Japanese R. solani isolates on Brachypodium distachyon and barley. Two isolates, AG-1 IA (from rice) and AG-4 HG-I+II (from cauliflower), infected leaves of both plants, but only AG-4 HG-I+II infected roots. B. distachyon accessions Bd3-1 and Gaz-4 and barley cultivar 'Morex' exhibited enhanced resistance to both isolates compared to B. distachyon Bd21 and barley cultivars 'Haruna Nijo' and 'Golden Promise'. During AG-1 IA infection, but not AG-4 HG-I+II infection, resistant Bd3-1 and Morex induced genes for salicylic acid (SA) and N-hydroxypipecolic acid (NHP) biosynthesis. Pretreatment with SA or NHP conferred resistance to AG-1 IA, but not AG-4 HG-I+II, in susceptible B. distachyon Bd21 and barley Haruna Nijo. On the leaves of susceptible Bd21 and Haruna Nijo, AG-1 IA developed extensive mycelial networks with numerous infection cushions, which are specialized infection structures well-characterized in rice sheath blight. In contrast, AG-4 HG-I+II formed dispersed mycelial masses associated with underlying necrosis. We propose that the R. solani species complex encompasses at least two distinct infection strategies: AG-1 IA exhibits a hemibiotrophic lifestyle, while AG-4 HG-I+II follows a predominantly necrotrophic strategy. No potential conflict of interest relevant to this article was reported.

Springer Science and Business Media LLC Acta Medica Okayama 0040-5752 137 9 2024 Mutations in starch BRANCHING ENZYME 2a suppress the traits caused by the loss of ISOAMYLASE1 in barley 212 EN Ryo Matsushima Institute of Plant Science and Resources, Okayama University Hiroshi Hisano Institute of Plant Science and Resources, Okayama University June-Sik Kim Institute of Plant Science and Resources, Okayama University Rose McNelly John Innes Centre, Norwich Research Park Naoko F. Oitome Department of Biological Production, Akita Prefectural University David Seung John Innes Centre, Norwich Research Park Naoko Fujita Department of Biological Production, Akita Prefectural University Kazuhiro Sato Institute of Plant Science and Resources, Okayama University The genetic interactions among starch biosynthesis genes can be exploited to alter starch properties, but they remain poorly understood due to the various combinations of mutations to be tested. Here, we isolated two novel barley mutants defective in starch BRANCHING ENZYME 2a (hvbe2a-1 and hvbe2a-2) based on the starch granule (SG) morphology. Both hvbe2a mutants showed elongated SGs in the endosperm and increased resistant starch content. hvbe2a-1 had a base change in HvBE2a gene, substituting the amino acid essential for its enzyme activity, while hvbe2a-2 is completely missing HvBE2a due to a chromosomal deletion. Further genetic crosses with barley isoamylase1 mutants (hvisa1) revealed that both hvbe2a mutations could suppress defects in endosperm caused by hvisa1, such as reduction in starch, increase in phytoglycogen, and changes in the glucan chain length distribution. Remarkably, hvbe2a mutations also transformed the endosperm SG morphology from the compound SG caused by hvisa1 to bimodal simple SGs, resembling that of wild-type barley. The suppressive impact was in competition with floury endosperm 6 mutation (hvflo6), which could enhance the phenotype of hvisa1 in the endosperm. In contrast, the compound SG formation induced by the hvflo6 hvisa1 mutation in pollen was not suppressed by hvbe2a mutations. Our findings provide new insights into genetic interactions in the starch biosynthetic pathway, demonstrating how specific genetic alterations can influence starch properties and SG morphology, with potential applications in cereal breeding for desired starch properties. No potential conflict of interest relevant to this article was reported.

Springer Science and Business Media LLC Acta Medica Okayama 0040-5752 136 4 2023 FLOURY ENDOSPERM 6 mutations enhance the sugary phenotype caused by the loss of ISOAMYLASE1 in barley 94 EN Ryo Matsushima Institute of Plant Science and Resources, Okayama University Hiroshi Hisano Institute of Plant Science and Resources, Okayama University Ivan Galis Institute of Plant Science and Resources, Okayama University Satoko Miura Department of Biological Production, Akita Prefectural University Naoko Crofts Department of Biological Production, Akita Prefectural University Yuto Takenaka College of Life Sciences, Ritsumeikan University Naoko F. Oitome Department of Biological Production, Akita Prefectural University Takeshi Ishimizu College of Life Sciences, Ritsumeikan University Naoko Fujita Department of Biological Production, Akita Prefectural University Kazuhiro Sato Institute of Plant Science and Resources, Okayama University Starch is a biologically and commercially important glucose polymer synthesized by plants as semicrystalline starch granules (SGs). Because SG morphology affects starch properties, mutants with altered SG morphology may be useful in breeding crops with desirable starch properties, including potentially novel properties. In this study, we employed a simple screen for mutants with altered SG morphology in barley (Hordeum vulgare). We isolated mutants that formed compound SGs together with the normal simple SGs in the endosperm and found that they were allelic mutants of the starch biosynthesis genes ISOAMYLASE1 (HvISA1) and FLOURY ENDOSPERM 6 (HvFLO6), encoding starch debranching enzyme and CARBOHYDRATE-BINDING MODULE 48-containing protein, respectively. We generated the hvflo6 hvisa1 double mutant and showed that it had significantly reduced starch biosynthesis and developed shrunken grains. In contrast to starch, soluble ƒ¿-glucan, phytoglycogen, and sugars accumulated to higher levels in the double mutant than in the single mutants. In addition, the double mutants showed defects in SG morphology in the endosperm and in the pollen. This novel genetic interaction suggests that hvflo6 acts as an enhancer of the sugary phenotype caused by hvisa1 mutation. No potential conflict of interest relevant to this article was reported.

Japanese Society of Breeding Acta Medica Okayama 1344-7610 71 4 2021 Targeted genome modifications in cereal crops 405 416 EN Hiroshi Hisano Institute of Plant Science and Resources, Okayama University Fumitaka Abe Institute of Crop Science, National Agriculture and Food Research Organization Robert E. Hoffie Leibniz Institute of Plant Genetics and Crop Plant Research (IPK) Jochen Kumlehn Leibniz Institute of Plant Genetics and Crop Plant Research (IPK) The recent advent of customizable endonucleases has led to remarkable advances in genetic engineering, as these molecular scissors allow for the targeted introduction of mutations or even precisely predefined genetic modifications into virtually any genomic target site of choice. Thanks to its unprecedented precision, efficiency, and functional versatility, this technology, commonly referred to as genome editing, has become an effective force not only in basic research devoted to the elucidation of gene function, but also for knowledgebased improvement of crop traits. Among the different platforms currently available for site-directed genome modifications, RNA-guided clustered regularly interspaced short palindromic repeats (CRISPR)-associated (Cas) endonucleases have proven to be the most powerful. This review provides an application-oriented overview of the development of customizable endonucleases, current approaches to cereal crop breeding, and future opportunities in this field. No potential conflict of interest relevant to this article was reported.

Wiley Acta Medica Okayama 1467-7644 2021 Regulation of germination by targeted mutagenesis of grain dormancy genes in barley 1 10 EN Hiroshi Hisano Institute of Plant Science and Resources, Okayama University Robert E. Hoffie Leibniz Institute of Plant Genetics and Crop Plant Research (IPK) Gatersleben Fumitaka Abe Institute of Crop Science, NARO Hiromi Munemori Institute of Plant Science and Resources, Okayama University Takakazu Matsuura Institute of Plant Science and Resources, Okayama University Masaki Endo Institute of Agrobiological Sciences, NARO Masafumi Mikami Institute of Agrobiological Sciences, NARO Shingo Nakamura Institute of Crop Science, NARO Jochen Kumlehn Leibniz Institute of Plant Genetics and Crop Plant Research (IPK) Gatersleben Kazuhiro Sato Institute of Plant Science and Resources, Okayama University High humidity during harvest season often causes pre-harvest sprouting in barley (Hordeum vulgare). Prolonged grain dormancy prevents pre-harvest sprouting; however, extended dormancy can interfere with malt production and uniform germination upon sowing. In this study, we used Cas9-induced targeted mutagenesis to create single and double mutants in QTL FOR SEED DORMANCY 1 (Qsd1) and Qsd2 in the same genetic background. We performed germination assays in independent qsd1 and qsd2 single mutants, as well as in two double mutants, which revealed a strong repression of germination in the mutants. These results demonstrated that normal early grain germination requires both Qsd1 and Qsd2 function. However, germination of qsd1 was promoted by treatment with 3% hydrogen peroxide, supporting the notion that the mutants exhibit delayed germination. Likewise, exposure to cold temperatures largely alleviated the block of germination in the single and double mutants. Notably, qsd1 mutants partially suppress the long dormancy phenotype of qsd2, while qsd2 mutant grains failed to germinate in the light, but not in the dark. Consistent with the delay in germination, abscisic acid accumulated in all mutants relative to the wild type, but abscisic acid levels cannot maintain long-term dormancy and only delay germination. Elucidation of mutant allele interactions, such as those shown in this study, are important for fine-tuning traits that will lead to the design of grain dormancy through combinations of mutant alleles. Thus, these mutants will provide the necessary germplasm to study grain dormancy and germination in barley. No potential conflict of interest relevant to this article was reported.

Frontiers Media SA Acta Medica Okayama 1664-302X 12 2021 Identification of a Novel Quinvirus in the Family Betaflexiviridae That Infects Winter Wheat 715545 EN Hideki Kondo Institute of Plant Science and Resources (IPSR), Okayama University Naoto Yoshida Agricultural Research Institute, HOKUREN Federation of Agricultural Cooperatives Miki Fujita Institute of Plant Science and Resources (IPSR), Okayama University Kazuyuki Maruyama Institute of Plant Science and Resources (IPSR), Okayama University Kiwamu Hyodo Institute of Plant Science and Resources (IPSR), Okayama University Hiroshi Hisano Institute of Plant Science and Resources (IPSR), Okayama University Tetsuo Tamada Institute of Plant Science and Resources (IPSR), Okayama University Ida Bagus Andika College of Plant Health and Medicine, Qingdao Agricultural University Nobuhiro Suzuki Institute of Plant Science and Resources (IPSR), Okayama University Yellow mosaic disease in winter wheat is usually attributed to the infection by bymoviruses or furoviruses; however, there is still limited information on whether other viral agents are also associated with this disease. To investigate the wheat viromes associated with yellow mosaic disease, we carried out de novo RNA sequencing (RNA-seq) analyses of symptomatic and asymptomatic wheat-leaf samples obtained from a field in Hokkaido, Japan, in 2018 and 2019. The analyses revealed the infection by a novel betaflexivirus, which tentatively named wheat virus Q (WVQ), together with wheat yellow mosaic virus (WYMV, a bymovirus) and northern cereal mosaic virus (a cytorhabdovirus). Basic local alignment search tool (BLAST) analyses showed that the WVQ strains (of which there are at least three) were related to the members of the genus Foveavirus in the subfamily Quinvirinae (family Betaflexiviridae). In the phylogenetic tree, they form a clade distant from that of the foveaviruses, suggesting that WVQ is a member of a novel genus in the Quinvirinae. Laboratory tests confirmed that WVQ, like WYMV, is potentially transmitted through the soil to wheat plants. WVQ was also found to infect rye plants grown in the same field. Moreover, WVQ-derived small interfering RNAs accumulated in the infected wheat plants, indicating that WVQ infection induces antiviral RNA silencing responses. Given its common coexistence with WYMV, the impact of WVQ infection on yellow mosaic disease in the field warrants detailed investigation. No potential conflict of interest relevant to this article was reported.

Cell Press Acta Medica Okayama 2666-1667 1 2 2020 Protocol for Genome Editing to Produce Multiple Mutants in Wheat 100053 EN Fumitaka Abe Division of Basic Research, Institute of Crop Science, NARO Yuji Ishida Plant Innovation Center, Japan Tobacco Inc. Hiroshi Hisano Institute of Plant Science and Resources, Okayama University Masaki Endo Division of Applied Genetics, Institute of Agrobiological Sciences, NARO Toshihiko Komari Plant Innovation Center, Japan Tobacco Inc. Seiichi Toki Division of Applied Genetics, Institute of Agrobiological Sciences, NARO Kazuhiro Sato Institute of Plant Science and Resources, Okayama University Here, we describe a protocol for producing multiple recessive mutants via genome editing in hexaploid wheat (Triticum aestivum) cv. Fielder. Using Agrobacterium-delivered CRISPR/Cas9 and three sub-genome-specific primer sets, all possible combinations of single, double, and triple transgene-free mutants can be generated. The technique for acceleration of generation advancement with embryo culture reduces time for mutant production. The mutants produced by this protocol can be used for the analysis of gene function and crop improvement. For complete details on the use and execution of this protocol, please refer to Abe et al. (2019). No potential conflict of interest relevant to this article was reported.

Frontiers Media Acta Medica Okayama 1664-302X 11 2020 Virome Analysis of Aphid Populations That Infest the Barley Field: The Discovery of Two Novel Groups of Nege/Kita-Like Viruses and Other Novel RNA Viruses 509 EN Hideki Kondo Institute of Plant Science and Resources (IPSR), Okayama University Miki Fujita Institute of Plant Science and Resources (IPSR), Okayama University Hiroshi Hisano Institute of Plant Science and Resources (IPSR), Okayama University Kiwamu Hyodo Institute of Plant Science and Resources (IPSR), Okayama University Ida Bagus Andika College of Plant Health and Medicine, Qingdao Agricultural University Nobuhiro Suzuki Institute of Plant Science and Resources (IPSR), Okayama University Aphids (order Hemiptera) are important insect pests of crops and are also vectors of many plant viruses. However, little is known about aphid-infecting viruses, particularly their diversity and relationship to plant viruses. To investigate the aphid viromes, we performed deep sequencing analyses of the aphid transcriptomes from infested barley plants in a field in Japan. We discovered virus-like sequences related to nege/kita-, flavi-, tombus-, phenui-, mononega-, narna-, chryso-, partiti-, and luteoviruses. Using RT-PCR and sequence analyses, we determined almost complete sequences of seven nege/kitavirus-like virus genomes; one of which was a variant of the Wuhan house centipede virus (WHCV-1). The other six seem to belong to four novel viruses distantly related to Wuhan insect virus 9 (WhIV-9) or Hubei nege-like virus 4 (HVLV-4). We designated the four viruses as barley aphid RNA virus 1 to 4 (BARV-1 to -4). Moreover, some nege/kitavirus-like sequences were found by searches on the transcriptome shotgun assembly (TSA) libraries of arthropods and plants. Phylogenetic analyses showed that BARV-1 forms a clade with WHCV-1 and HVLV-4, whereas BARV-2 to -4 clustered with WhIV-9 and an aphid virus, Aphis glycines virus 3. Both virus groups (tentatively designated as Centivirus and Aphiglyvirus, respectively), together with arthropod virus-like TSAs, fill the phylogenetic gaps between the negeviruses and kitaviruses lineages. We also characterized the flavi/jingmen-like and tombus-like virus sequences as well as other RNA viruses, including six putative novel viruses, designated as barley aphid RNA viruses 5 to 10. Interestingly, we also discovered that some aphid-associated viruses, including nege/kita-like viruses, were present in different aphid species, raising a speculation that these viruses might be distributed across different aphid species with plants being the reservoirs. This study provides novel information on the diversity and spread of nege/kitavirus-related viruses and other RNA viruses that are associated with aphids. No potential conflict of interest relevant to this article was reported.

Nature Publishing Group Acta Medica Okayama 2045-2322 9 2019 Imaging Amyloplasts in the Developing Endosperm of Barley and Rice 3745 EN Ryo Matsushima Institute of Plant Science and Resources, Okayama University Hiroshi Hisano Institute of Plant Science and Resources, Okayama University Amyloplasts are plant-specific organelles responsible for starch biosynthesis and storage. Inside amyloplasts, starch forms insoluble particles, referred to as starch grains (SGs). SG morphology differs between species and SG morphology is particularly diverse in the endosperm of Poaceae plants, such as rice (Oryza sativa) and barley (Hordeum vulgare), which form compound SGs and simple SGs, respectively. SG morphology has been extensively imaged, but the comparative imaging of amyloplast morphology has been limited. In this study, SG-containing amyloplasts in the developing endosperm were visualized using stable transgenic barley and rice lines expressing amyloplast stroma-targeted green fluorescent protein fused to the transit peptide (TP) of granule-bound starch synthase I (TP-GFP). The TP-GFP barley and rice plants had elongated amyloplasts containing multiple SGs, with constrictions between the SGs. In barley, some amyloplasts were connected by narrow protrusions extending from their surfaces. Transgenic rice lines producing amyloplast membrane-localized SUBSTANDARD STARCH GRAIN6 (SSG6)-GFP were used to demonstrate that the developing amyloplasts contained multiple compound SGs. TP-GFP barley can be used to visualize the chloroplasts in leaves and other plastids in pollen and root in addition to the endosperm, therefore it provides as a useful tool to observe diverse plastids. No potential conflict of interest relevant to this article was reported.