TR75, a rice (Oryza sativa L. var. Sasanishikj) mutant resistant to 5-methyltryptophan (5MT) was segregated from the progenies of its initial mutant line, TR1. The 5MT resistance of TR75 was inherited in the M$_{8}$ generations as a single dominant nuclear gene, and was also expressed in callus derived from seeds, roots, and anthers as well as in the seedlings. The callus induced from these organs could grow at 50 mg/1 of 5MT, whereas the growth of wild-type callus was completely inhibited even at 25 mg/1. The seedlings of TR75 did not show resistance to L-azetidine-2-carboxylic acid, S-2-aminoethyl-L-cysteine, p-fluoro-DL-phenylalanine. The content of free amino acids in the TR75 homozygous seeds increased approximately 1.5 to 2.0 fold compared to wild-type seeds. Especially, the contents of tryptophan, phenylalanine and aspartic acid were 5.0, 5.3 and 2.7 times higher than those of wild-type seeds, respectively.y.
Proteomic analysis was performed in order to identify proteomic changes by salt stress between the Japonica cv. Donganbyeo (WT) and two salt-tolerant (ST) mutant lines by using the SDS-PAGE and 2-DE. Two salt tolerant rice mutant lines, ST-87 and ST-301, were selected by in vitro mutagenesis with gamma-ray. Three-week-old seedlings were treated with 171 mM NaCl for 7 days. In the SDS-PAGE, three proteins with molecular weights of 27, 46 and 58 kDa were highly increased under salt treatment. Total proteins from shoots of both WT and ST-lines were separated by two-dimensional gel electrophoresis. In 2-DE, 201, 226, 217 and 213 protein spots were detected in the untreated-or treated-WT and untreated- or treated-ST-87, respectively. Of theses, 17 and 10 protein spots were up- and down-regulated under salt stress in the WT, respectively. While, 16 and 8 protein spots were up- and down-regulated under salt stress in the ST-87, respectively, compared with the untreated plants. High intensity or de novo synthesized proteins were analyzed by MALDI-TOF/MS analysis.
Kang, Beom-Ryong;Han, Song-Hee;Zdor, Rob E.;Anderson, Anne J.;Spencer, Matt;Yang, Kwang-Yeol;Kim, Yong-Hwan;Lee, Myung-Chul;Cho, Baik-Ho;Kim, Young-Cheol
Journal of Microbiology and Biotechnology
/
v.17
no.4
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pp.586-593
/
2007
Seed coating by a phenazine-producing bacterium, Pseudomonas chlororaphis O6, induced dose-dependent inhibition of germination in wheat and barley seeds, but did not inhibit germination of rice or cucumber seeds. In wheat seedlings grown from inoculated seeds, phenazine production levels near the seed were higher than in the roots. Deletion of the gacS gene reduced transcription from the genes required for phenazine synthesis, the regulatory phzI gene and the biosynthetic phzA gene. The inhibition of seed germination and the induction of systemic disease resistance against a bacterial soft-rot pathogen, Erwinia carotovora subsp. carotovora, were impaired in the gacS and phzA mutants of P chlororaphis O6. Culture filtrates of the gacS and phzA mutants of P. chlororaphis O6 did not inhibit seed germination of wheat, whereas that of the wild-type was inhibitory. Our results showed that the production of phenazines by P. chlororaphis O6 was correlated with reduced germination of barley and wheat seeds, and the level of systemic resistance in tobacco against E. carotovora.
Kim, Sunyoung;Park, Jungwook;Kim, Ji Hyeon;Lee, Jongyun;Bang, Bongjun;Hwang, Ingyu;Seo, Young-Su
The Plant Pathology Journal
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v.29
no.3
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pp.249-259
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2013
Burkholderia glumae causes rice grain rot and sheath rot by producing toxoflavin, the expression of which is regulated by quorum sensing (QS). The QS systems of B. glumae rely on N-octanoyl homoserine lactone, synthesized by TofI and its cognate receptor TofR, to activate the genes for toxoflavin biosynthesis and an IclR-type transcriptional regulator gene, qsmR. To understand genome-wide transcriptional profiling of QS signaling, we employed RNAseq of the wild-type B. glumae BGR1 with QS-defective mutant, BGS2 (BGR1 tofI::${\Omega}$) and QS-dependent transcriptional regulator mutant, BGS9 (BGR1 qsmR::${\Omega}$). A comparison of gene expression profiling among the wild-type BGR1 and the two mutants before and after QS onset as well as gene ontology (GO) enrichment analysis from differential expressed genes (DEGs) revealed that genes involved in motility were highly enriched in TofI-dependent DEGs, whereas genes for transport and DNA polymerase were highly enriched in QsmR-dependent DEGs. Further, a combination of pathways with these DEGs and phenotype analysis of mutants pointed to a couple of metabolic processes, which are dependent on QS in B. glumae, that were directly or indirectly related with bacterial motility. The consistency of observed bacterial phenotypes with GOs or metabolic pathways in QS-regulated genes implied that integration RNAseq with GO enrichment or pathways would be useful to study bacterial physiology and phenotypes.
Optimal conditions of producing red pigment by Monascus anka, Nakazawa, et Sato, IFO 4478 and 6540 were found to be at pH 6.0 and $30^{\circ}C$ for 10 days. When 3.0% steamed rice and 1.0% defatted sesame exfracts were used as substrates, the highest production of red pigment was yielded. Furthermore, mutants such as Monascus anka 4475-27 and 6540-185 induced by N -methyl-N-nitro-N-nitrosoguanidine (MNNG) treatment were found to produce pigment much more than parent strains.
Kim, Hong-Sik;Kim, Dea-Wook;Kim, Sun-Lim;Baek, Seong-Bum;Park, Hyoung-Ho;Hwang, Jong-Jin;Kim, Si-Ju
Korean Journal of Breeding Science
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v.43
no.5
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pp.375-382
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2011
A chemical, MNU-induced hulless barley mutant line designated as 'Mutant 98 (M98)' was developed from a Korean hulless waxy barley cultivar, 'Chalssalbori'. The objective of the study was to determine the genetic basis of 'M98' and the possibility of using 'M98' as breeding parent to improve lysine level. Compared to 'Chalssalbori', 'M98' had large embryo and higher lysine content in both the embryo and endosperm. Significantly different lysine content in 'M98' and the other high-lysine barley mutant stocks was observed for two years. However, the genotype by year interaction was not significant. 'M98' was higher than the other high-lysine barley mutant stocks in the percentage of lysine of total amino acid composition (0.75%). The trait of shrunken endosperm of 'M98', which was typical in the high-lysine mutants, was inherited by a single recessive gene. Based on seed morphology and lysine content of $F_1$ seeds, 'M98' had a genetically different gene from the other high-lysine mutants for shrunken endosperm. Segregation of $F_2$ for plump/shrunken endosperm did not fit the expected ratio of Mendelian inheritance except for only one cross combination (GSHO1784 (lys1)/M98). The amino acid analysis of $F_5$ and $F_6$ progenies from the cross between 'M98' and 'Chalssalbori' revealed that the attempt to increase the range of lysine content of plump lines did not go beyond the limit of the average high-lysine barley germplasm.
It is crucial to develop a miniaturized cultivation method for large and rapid screening of high-yielding mutants of monacolin-K, a powerful anti-hypercholesterolemic secondary metabolite biosynthesized by the fungal cells of Monascus ruber. In order to investigate as many strains as possible in a short time, a miniaturized fermentation method especially suitable for the cultivation of the filamentous Monascus mutants was developed using $50m{\ell}$ culture-tube ($7m{\ell}$ of working volume) instead of the traditional $250m{\ell}$ flask ($50m{\ell}$ of working volume). Generally, in filamentous fungal cell fermentations, morphologies in growth and production cultures should be maintained as thick filamentous and compact-pelleted (usually less than 1 mm in diameter) forms, respectively, for enhanced production of secondary metabolites in final production cultures. In this study, we intended to induce the respective optimal morphologies in the miniaturized culture system for the purpose of rapid screening of overproducers. Miniaturized growth culture system was successfully developed due to the mass production of spores in the statistically optimized solid medium. When large amounts of spores were inoculated into the growth cultures, and brown rice flour (20 g/L) was also supplemented to the growth medium, dense filamentous morphologies were successfully induced in the growth cultures performed with the 50 ml culture tubes. It was implied that the amounts of spores inoculated into the growth tube-cultures and the growth medium components should be the key factors for the induction of the filamentous forms in the growth fermentations. Furthermore, in order to statistically optimize production medium, multiple experiments based on Plackett-Burman design and response surface method (RSM) were carried out, resulting in more than 2 fold enhanced production of monacolin-K in the final production cultures with the optimized production medium. Notably, under the production culture conditions with the statistically optimized medium, optimal pellet sizes below 1 mm in diameter were reproducibly induced, in contrast to the thick and viscous filamentous morphologies observed in the previous production cultures.
Fusarium graminearum (teleomorph Gibberella zeae) is an important plant pathogen that causes head blight of major cereal crops such as wheat, barley, and rice, as well as causing ear and stalk rot on maize worldwide. Plant diseases caused by this fungus lead to severe yield losses and accumulation of harmful mycotoxins in infected cereals [1]. Fungi utilize spore production as a mean to rapidly avoid unfavorable environmental conditions and to amplify their population. Spores are produced sexually and asexually and their production is precisely controlled. Upstream developmental activators consist of fluffy genes have been known to orchestrate early induction of condiogenesis in a model filamentous fungus Aspergillus nidulans. To understand the molecular mechanisms underlying conidiogenesis in F. graminearum, we characterized functions of the F. graminearum fluffy gene homologs [2]. We found that FlbD is conserved regulatory function for conidiogenesis in both A. nidulans and F. graminearum among five fluffy gene homologs. flbD deletion abolished conidia and perithecia production, suggesting that FlbD have global roles in hyphal differentiation processes in F. graminearum. We further identified and functionally characterized the ortholog of AbaA, which is involved in differentiation from vegetative hyphae to conidia and known to be absent in F. graminearum [3]. Deletion of abaA did not affect vegetative growth, sexual development, or virulence, but conidium production was completely abolished and thin hyphae grew from abnormally shaped phialides in abaA deletion mutants. Overexpression of abaA resulted in pleiotropic defects such as impaired sexual and asexual development, retarded conidium germination, and reduced trichothecene production. AbaA localized to the nuclei of phialides and terminal cells of mature conidia. Successful interspecies complementation using A. nidulans AbaA and the conserved AbaA-WetA pathway demonstrated that the molecular mechanisms responsible for AbaA activity are conserved in F. graminearum as they are in A. nidulans. F. graminearum ortholog of Aspergillus nidulans wetA has been shown to be involved in conidiogenesis and conidium maturation [4]. Deletion of F. graminearum wetA did not alter mycelial growth, sexual development, or virulence, but the wetA deletion mutants produced longer conidia with fewer septa, and the conidia were sensitive to acute stresses, such as oxidative stress and heat stress. Furthermore, the survival rate of aged conidia from the F. graminearum wetA deletion mutants was reduced. The wetA deletion resulted in vigorous generation of single-celled conidia through autophagy-dependent microcycle conidiation, indicating that WetA functions to maintain conidia dormancy by suppressing microcycle conidiation in F. graminearum. In A. nidulans, FlbB physically interacts with FlbD and FlbE, and the resulting FlbB/FlbE and FlbB/FlbD complexes induce the expression of flbD and brlA, respectively. BrlA is an activator of the AbaA-WetA pathway. AbaA and WetA are required for phialide formation and conidia maturation, respectively [5]. In F. graminearum, the AbaA-WetA pathway is similar to that of A. nidulans, except a brlA ortholog does not exist. Amongst the fluffy genes, only fgflbD has a conserved role for regulation of the AbaA-WetA pathway.
Li Zhang;Dong Li;Min Lu;Zechi Wu;Chaotian Liu;Yingying Shi;Mengyu Zhang;Zhangjie Nan;Weixiang Wang
The Plant Pathology Journal
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v.39
no.4
/
pp.361-373
/
2023
In plant-pathogen interactions, Magnaporthe oryzae causes blast disease on more than 50 species of 14 monocot plants, including important crops such as rice, millet, and most 15 recently wheat. M. oryzae is a model fungus for studying plant-microbe interaction, and the main source for fungal pathogenesis in the field. Here we report that MoJMJD6 is required for conidium germination and appressorium formation in M. oryzae. We obtained MoJMJD6 mutants (ΔMojmjd6) using a target gene replacement strategy. The MoJMD6 deletion mutants were delayed for conidium germination, glycogen, and lipid droplets utilization and consequently had decreased virulence. In the ΔMojmjd6 null mutants, global histone methyltransferase modifications (H3K4me3, H3K9me3, H3K27me3, and H3K36me2/3) of the genome were unaffected. Taken together, our results indicated that MoJMJD6 function as a nuclear protein which plays an important role in conidium germination and appressorium formation in the M. oryzae. Our work provides insights into MoJMJD6-mediated regulation in the early stage of pathogenesis in plant fungi.
The endeavors enhancing the grain quality of high-yielding japonica rice were steadily continued during 1980s-1990s along with the self-sufficiency of rice production and the increasing demands of high-quality rices. During this time, considerably great progress and success was obtained in development of high-quality japonica cultivars and quality evaluation techniques including the elucidation of interrelationship between the physicochemical properties of rice grain and the physical or palatability components of cooked rice. In 1990s, some high-quality japonica rice cultivars and special rices adaptable for food processing such as large kernel, chalky endosperm, aromatic and colored rices were developed and its objective preference and utility was also examined by a palatability meter, rapid-visco analyzer and texture analyzer, Recently, new special rices such as extremely low-amylose dull or opaque non-glutinous endosperm mutants were developed. Also, a high-lysine rice variety was developed for higher nutritional utility. The water uptake rate and the maximum water absorption ratio showed significantly negative correlations with the K/Mg ratio and alkali digestion value(ADV) of milled rice. The rice materials showing the higher amount of hot water absorption exhibited the larger volume expansion of cooked rice. The harder rices with lower moisture content revealed the higher rate of water uptake at twenty minutes after soaking and the higher ratio of maximum water uptake under the room temperature condition. These water uptake characteristics were not associated with the protein and amylose contents of milled rice and the palatability of cooked rice. The water/rice ratio (in w/w basis) for optimum cooking was averaged to 1.52 in dry milled rices (12% wet basis) with varietal range from 1.45 to 1.61 and the expansion ratio of milled rice after proper boiling was average to 2.63(in v/v basis). The major physicochemical components of rice grain associated with the palatability of cooked rice were examined using japonica rice materials showing narrow varietal variation in grain size and shape, alkali digestibility, gel consistency, amylose and protein contents, but considerable difference in appearance and texture of cooked rice. The glossiness or gross palatability score of cooked rice were closely associated with the peak, hot paste and consistency viscosities of viscosities with year difference. The high-quality rice variety "IIpumbyeo" showed less portion of amylose on the outer layer of milled rice grain and less and slower change in iodine blue value of extracted paste during twenty minutes of boiling. This highly palatable rice also exhibited very fine net structure in outer layer and fine-spongy and well-swollen shape of gelatinized starch granules in inner layer and core of cooked rice kernel compared with the poor palatable rice through image of scanning electronic microscope. Gross sensory score of cooked rice could be estimated by multiple linear regression formula, deduced from relationship between rice quality components mentioned above and eating quality of cooked rice, with high probability of determination. The $\alpha$-amylose-iodine method was adopted for checking the varietal difference in retrogradation of cooked rice. The rice cultivars revealing the relatively slow retrogradation in aged cooked rice were IIpumbyeo, Chucheongyeo, Sasanishiki, Jinbubyeo and Koshihikari. A Tonsil-type rice, Taebaegbyeo, and a japonica cultivar, Seomjinbyeo, showed the relatively fast deterioration of cooked rice. Generally, the better rice cultivars in eating quality of cooked rice showed less retrogradation and much sponginess in cooled cooked rice. Also, the rice varieties exhibiting less retrogradation in cooled cooked rice revealed higher hot viscosity and lower cool viscosity of rice flour in amylogram. The sponginess of cooled cooked rice was closely associated with magnesium content and volume expansion of cooked rice. The hardness-changed ratio of cooked rice by cooling was negatively correlated with solids amount extracted during boiling and volume expansion of cooked rice. The major physicochemical properties of rice grain closely related to the palatability of cooked rice may be directly or indirectly associated with the retrogradation characteristics of cooked rice. The softer gel consistency and lower amylose content in milled rice revealed the higher ratio of popped rice and larger bulk density of popping. The stronger hardness of rice grain showed relatively higher ratio of popping and the more chalky or less translucent rice exhibited the lower ratio of intact popped brown rice. The potassium and magnesium contents of milled rice were negatively associated with gross score of noodle making mixed with wheat flour in half and the better rice for noodle making revealed relatively less amount of solid extraction during boiling. The more volume expansion of batters for making brown rice bread resulted the better loaf formation and more springiness in rice breed. The higher protein rices produced relatively the more moist white rice bread. The springiness of rice bread was also significantly correlated with high amylose content and hard gel consistency. The completely chalky and large grain rices showed better suitability far fermentation and brewing. The glutinous rice were classified into nine different varietal groups based on various physicochemical and structural characteristics of endosperm. There was some close associations among these grain properties and large varietal difference in suitability to various traditional food processing. Our breeding efforts on improvement of rice quality for high palatability and processing utility or value-adding products in the future should focus on not only continuous enhancement of marketing and eating qualities but also the diversification in morphological, physicochemical and nutritional characteristics of rice grain suitable for processing various value-added rice foods.ice foods.
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