To evaluate expression level of HMW-GS protein qualitatively and quantitatively, we separated glutenin fractions and conducted two-dimensional electrophoresis (2DE) in 32 cultivars of Korean wheat for the use of as the basis of wheat breeding. The average spot number of HMW-GS in all Korean wheat cultivars was 11.78 which included 1.31, 5.53 and 4.94 to Glu-A1, Glu-B1 and Glu-D1 loci, respectively. Cultivars harboring 1, 2* subunits had many spots more than ones harboring null allele in Glu-A1 loci because there is no difference of spots between Glu-B1 and Glu-D1 loci. In total spot number of HMW-GS, the highest one was Jokyung as 18 and Dahong the lowest as 7. When the Korean wheat cultivars were compared with the Chinese spring in the average relative expression level, Korean one’s were lower as 0.44. Especially, Gobun was the highest as 1.11 and Eunpa was the lowest as 0.24. Also we investigated phylogenetic relationship based on both frequency of HMW-GS spots and quantification value of each spot to all HMW-GS spots. As a result, Korean the varieties of Korean wheat could be classified into six groups.

LMW-GSs represent approximately 1/3 of the total wheat gluten fraction, which have not been widely studied, even though they are important in the context of wheat end-use quality. In this study, we report on the qualitative and quantitative analysis of LMW-GS in korean wheat cultivars by 2DE in 32 cultivars of Korean wheat for the use of the basis of wheat breeding. We firstly identified spots corresponding each of Glu-3 alleles. The 2DE results for each cultivar will be used as reference map or protein marker discriminating wheat cultivars, wheat and rice, imported and Korean flour. Unexpectedly, five LMW-GS spots were found to be expressed at a common position in hexaploid wheat cultivars, and these spots might play something in glutenin biosynthesis. Total spot numbers were expressed variously between 20 and 10, and average spot number was shown 17.12. The average number of spots in Glu-A3, Glu-B3 and Glu-D3 were 3.0, 4.56 and 2.96 respectively. When the Korean wheat cultivars were compared with the Chinese spring (1.0) in the average relative expression level, Korean one’s were lower as 0.67. Especially, Gobun was the highest as 1.32 and Baekjoong was the lowest as 0.24. Also we investigated phylogenetic relationship based on frequency of HMW-GS spots and quantification value of each spot to all LMW-GS spots. As a result, the varieties of Korean wheat could be classified into five groups.

The THO/TREX complex mediates the transport of nascent mRNAs from the nucleus towards the cytoplasm in animals, and it has a role in small RNA-dependent processes in plants. Here we describe five mutant alleles of Arabidopsis thaliana THO2, whichencodes a core subunit of the plant THO/TREX complex. tho2 mutants present strong developmental defects resembling those in plants compromised in microRNA (miRNA) activity. In agreement, not only the levels of siRNAs, but also of mature miRNAs were reduced in tho2 mutants. As a consequence miRNA target mRNAs accumulated to higher levels than in wild type. Yeast two hybrid experiments showed that THO2 does not seem to interact with any of the known miRNA biogenesis components, implying a more indirect role of THOs in small RNA biogenesis. We also detected alterations in the splicing pattern of genes encoding Serine/Arginine-rich proteins in tho2 mutants, suggesting a previously unappreciated role of the THO/TREX complex in alternative splicing.

Gibberellic acid (GA) is a well-characterized plant hormone, which plays a critical role in various plant growth and development. including stem elongation, floral indcution and seed development. GA is known to cause enlargement of ripening fruits and, especially in grapevines, GA shows a unique function: the induction of seedlessness in seeded grape varieties. However, despite extensive previous studies about GA, there has been no clear verification of the mechanism that induces seedlessness in grapes. To understand how GA treatment results in artificial parthenocarpy of seeded grapes at molecular levels, we analyzed transcriptional changes in seeded grapes with and without GA application in various inflorescence developmental stages using RNA-seq. At 14 days before flowering (DBF), seeded grapes were treated with 100 ppm GA and clusters were collected at three developmental stages: 7 DBF, full bloom, and 5 days after flowering (DAF). Of a total of 28,974 genes that were mapped to grape genome reference sequences, 7,013 and 9,064 genes were up- and down-regulated, respectively, in the GA-treated grape as compared to the non-GA-treated control at 7 DBF, full bloom, and 5 DAF. Clustering analysis revealed that these genes could be grouped into 9 clusters with different expression patterns. We also carried out functional annotation based on gene ontology categories. There were significant differences in the expression of the GA and auxin-related gene families. These findings expand our understanding of the complex molecular and cellular mechanisms of GA-induced parthenocarpy of grapes and provide a foundation for future studies on seed development in grapevines.

Phosphorus is one of the macronutrients essential for plant growth and development, as well as crop productivity. Many soils around the world are deficient in phosphate (Pi) that plants can utilize. To cope with the stress of Pi starvation, plants have evolved many adaptive strategies, such as changes of root architecture and enhanced Pi acquisition form soil. To understand molecular mechanism underlying Pi starvation stress signaling, we characterized the activation-tagged mutant showing altered responses to Pi deficiency compared to wild type Arabidopsis and named hsp3 (hypersensitive to Pi starvation3). hsp3 mutant exhibits enhanced phosphate transporter activity, resulting in higher Pi content than wild type. However, in root architectural change under Pi starvation, hsp3 shows hyposensitive responses than wild type, such as longer primary root elongation, lower lateral root density. Histochemical analysis using hsp3 mutant expressing auxin-responsive DR5::GUS reporter gene, indicated that auxin allocation from primary to lateral roots under Pi starvation is aborted in hsp3 mutant. Molecular genetic analysis of hsp3 mutant revealed that the mutant phenotype is caused by the lesion in ENHANCED SILENCING PHENOTYPE4 (ESP4) gene whose function is proposed in mRNA 3’ end processing. Here, we propose that mRNA processing plays a crucial role in Pi homeostasis in Arabidopsis.

Many viruses infect cucurbits. One of the well-known symptoms is mosaic disease. Those that cause mosaic are cucumber mosaic virus (CMV), squash mosaic virus (SqMV), watermelon mosaic virus (WMV), zucchini yellow mosaic virus (ZYMV) and cucumber green mottle mosaic virus (CGMMV). WMV resistant GM squash was developed many years ago in the United States and it was on the market, but no further information was available by now pertinent to commercial aspect. Usually these viruses are not easily controlled by frequent applications of chemicals that target the insect as carriers of viruses. Therefore, it is necessary to develop commercial varieties possessing resistance against viral diseases. We have developed GM watermelon rootstocks called gongdae, using a coat protein gene of CGMMV as transgene. Those GM watermelon rootstocks showed highly resistant to CGMMV, and have been crossed to get the several BC and T generation. In order to obtain the virus resistant watermelon, watermelon lines were crossed to the selected GM watermelon rootstock. Here, we present the successful watermelon cultivars that show resistance to CGMMV. The resistance must have obtained by transferring the transgene from the GM watermelon rootstock to watermelon line.

Drought tolerance is the ability of a plant to live, grow, and reproduce properly with limited water supply or under periodic conditions of water deficit. However, the climate changes and worldwide water shortages would result in the loss of applied water to irrigated land, increasing soil water deficit. To control the situation, we have carried out the international joint research project for the aim of developing that drought tolerance common wheat and durum wheat in Korea and Tunisia. Total 79 (41 common wheat, 39 durum wheat) Tunisian lines and 33 Korean wheat cultivars were incorporated in this study. Drought stress was applied for 25 days of stopping irrigation from the 3-leaf stage followed by re-watering for restoration in greenhouse. We selected top 13 (5 Korean line, 8 Tunisian line) tolerant lines and 11 (5 Korean, 6 Tunisian) susceptible lines based on growth parameter analysis. Primers (Operon primers and wheat Dreb1 gene) that have been known to be related drought resistance were applied to explain selected population. The correlation between PCR-based length polymorphism of selected lines and their resistance were evaluated. The obtained primer information will aid selection for drought tolerance durum as well as hexaploid common wheat.

Rice genetic resources are composed of various species and ecotypes, and each accession reveals different genetic and phenotypic characters. For the managenment of diverse rice genetic resources, seed integrity is important factor in that the individuals of one accession in self-pollinating crop might be homogeneous. To elevate the management efficiency of rice germplasm contrary to the phenotypic distinction, we focused on applicable microsatellite markers because this markers are widly used for genetic evaluation in diverse genetic resources with a character of high reproducibility and polymorphism. In this regard, we selected microsatellite markers based on genotypes; diversity set including 150 accessions using 249 SSR markers. As SSR loci with high PIC(polymorphism information content) values usually revealed multi bands in one accession, proper genotyping were difficult in these loci. Therefor, we checked the band clarity in addition to PIC values and chose 12 and 6 SSR markers finally. All accessions of rice diversity set were distinguished with the first marker set comprising 12 SSR markers, and only 3 combinations of tested accessions(0.03%, 3/11,175) showed same genotype with second marker set comprising 6 SSR markers. The tested 142 Korean bred varieties revealed 0.19%(19/10,011 combinations) and 0.69%(69/10,011 combinations) genotypic identity using first and second marker set, respectively. These newly selected markers might be useful for the analysis of genetic homogeneity and relationship in rice genetic resources.

As the drought is getting worse, Lot of studies related to drought stress in plant have been conducted. Recently whole genome sequencing of Brassica rapa ssp which is important vegetable crop to East Asians has been completed to enable Omic research. It is known that the drought damages occur in the early stage of plant development. Here, we performed shotgun proteomics analysis of B. rapa to observe the morphological characters, monitor the expression patterns of the identified proteins during drought stress, and detect the proteins related to drought stress. The three week old B. rapa grown in density of single plant in a single pot were used. Drought stress were treated as that a single plant in soil was removed from the pot and the plant with soil was exposed to air and light without watering. Leaves were immediately harvested before drought treatment, 24hr after drought treatment, and 48hr after drought treatment. The protein expression patterns were monitored by a quantitative shotgun proteomics analysis. Extracted proteins were separated in 1D-SDS-PAGE then the gel sliced into seven pieces. Chopped gels were ingel-digested. Peptides were assigned to mass spectrometry (Q-Exactive). The ms/ms spectra were analyzed through Proteome Discoverer. By combining all of the identified proteins in the seven sliced gel samples, total B. rapa proteome reference map was completed. Protein expression patterns were investigated by comparing the quantity of protein. With shotgun proteomic approach, we evaluated the changes in the quantity and finally discovered the candidate proteins related with drought stress.

We investigated whether sound could alter gene expression in plants. Using a sound-treated subtractive library, a set of sound-responsive genes in plants was demonstrated through mRNA expression analyses. Of them, the rbcS and ald genes, which are light responsive, up-regulated their expression with sound treatment in both light and in dark conditions. This suggested that sound could be used as a gene regulator instead of light. When we analyzed ald gene expression using various single wavelengths, a significant increase in mRNA levels was found at 125 or 250 Hz but decreased at 50 Hz, indicating that the gene responded to sound in a wavelength-specific manner. To determine whether the ald promoter respond to sound, we generated transgenic rice plants harboring the chimeric gene consisting of a 1,506-bp promoter fragment of the ald gene fused to Escherichia coli GUS reporter gene. Analyses of mRNA expressison level of three independent transgenic lines sound-treated with 50 or 250 Hz for 4 h showed that the Gus gene expression in all three transgenic lines was up regulated by 250 Hz, but down regulated by 50 Hz. These results correlated with sound responsive mRNA expression pattern observed for the ald gene in rice plants, indicating that the 1,506-bp ald promoter confers sound-responsiveness on a reporter gene in transgenic rice plants. We also investigated whether sound waves could improve salt tolerance in rice seedling. The rice seedlings were sound treated with 800 Hz for 1hr, and then treated with 0, 75, 150, and 225mM NaCl for 3 days to observe changes in physiological and morphological aspects. Sound treatment seedlings resulted in enhanced salt stress tolerance, mainly demonstrated by the sound treated seedlings exhibiting of increased root relative water contents (RWC), root length and weight, photochemical efficiency (ratio of variable to maximum fluorescence, Fv/Fm), and germination rate under salt stress condition. This demonstrates that a specific sound wave might be used, not only to alter gene expression in plant, but also to improve salt stress tolerance.

본 연구는 강원도 농업기술원 옥수수연구소에서 튀김옥수수 품종개발을 위하여 육성한 79개의 자식계통들에 대하여 SSR 분자마커를 이용하여 집단구조 및 association mapping 분석을 실시하였다. 집단구조 분석결과, 79개의 튀김옥수수 자식계통들은 groups I, II, III, IV, admixed group으로 구분되었다. 4개의 옥수수 자식계통은 group I에 포함되었고, Group II는 총 17개의 자식계통들이 포함되었다. 그리고 6개의 자식계통들은 Group III에 포함되었으며, 22개의 자식계통들은 Group IV에 포함되었다. 그리고 admixed group에는 30개 옥수수 자식계통들로 구성되었다. 더욱이 본 연구에서는 튀김옥수수 자식계통들에서 분석에 이용된 50개 SSR 마커와 13개의 질적, 양적 형질과의 연관성을 분석하기 위해서 association mapping 분석을 실시하였으며, false positive associations을 최소화하기 위해서 population structure(Q), kinship(K) 값을 이용하여 Q GLM과 Q+K MLM 분석을 실시하였다. 0.05의 유의수준에서, Q GLM 분석을 이용하여 총 44개의 SSR 마커가 12개의 형질과 association을 보였고, Q+K MLM을 이용하여 분석하였을 때, 총 25개의 SSR 마커가 12개의 형질과 association을 보였다. 그리고 0.01의 유의수준에서, Q GLM 분석에서는 34개의 SSR 마커와 12개 형질의 association을 확인하였다. 더욱이 Q+K MLM을 이용하여 8개의 SSR 마커는 5개의 형질과 association을 확인하였다. 본 연구에서 79개의 튀김옥수수 자식계통들에 대한 집단구조 및 association mapping 분석의 결과는 앞으로 강원도농업기술원 옥수수연구소에서 튀김옥수수 품종개발을 위한 계통 육성 및 교배조합 구성 등에 유용한 정보를 제공할 것으로 기대한다.

The molecular processing of upstream regulation of Pi response genes during Pi starvation remains inadequately understood. Several transcription factor have been studied that appear to regulate subsets of the responses to Pi stress either positively or negatively. MYB genes are responsive to one or multiple type of hormone and stress treatments. In this study, cDNA of the MYB have been cloned, and we generated Rice overexpressing plants for characterization of these genes. OsMYB gene function focused on phosphate conditions with rice and Arabidopsis transgenic plants. We selected 30 - T1 transgenic lines from T0 transgenic rices. those are shown high Pi content. The Pi contents of shoots part of transgenic plants were shown 10~20% increased Pi contents than WT, whereas roots have 30% increased Pi contents. As a result, OsMYB genes affect Pi uptake in plants. To investigate interactions between MYB proteins and phosphate signaling related genes. We demonstrate that Myb-binding sites (MBSs) exist in putative promoter of OsPT transporter by analysis of bioinformatics, and its bind specific MYB transcription factor. OsMYB expression is induced by low Pi and Pi deficiency, and its overexpression plants are shown morphological phenotype in Pi stress.

Excessive water stress can cause severe damage to sorghum and results in significant yield reduction. The aim of this study is to identify quantitative trait loci (QTL) for excessive water stress in sorghum. As a first step, two out of 21 bmr mutants were selected for their superior agronomic performance and Chlorophyll a fluorescence OJIP transient, and were crossed with an elite Korean cultivar, Hwangkeumchal, to construct mapping populations. One hundred ten out of 236 SSR primers showed polymorphism between two parens, which cover ten chromosomes of sorghum from different published SSR linkage maps of sorghum. Development of recombinant inbred lines from the crosses ‘25M2-0698 x Hwangkeumchal’ and ‘25M2-0404 x Hwangkeumchal’ are in progress using the single seed descendent method for generation acceleration.

Over the last decades, increasing natural disasters and climate change are considered as the major environmental problems facing the globe. Numerous studies have been indicated it would cause huge losses on agriculture, especially in the grain productivity. Therefore, several alternatives are suggested for boosting up productivity of wheat as one of the main human food crop. One of important strategy is proper management of inflorescence development and DELLA proteins have been elucidated to play pivotal roles in growth of many plant organs. In this study, putative negative regulator of DELLA protein, GAST (Gibberellic acids stimulated transcript) have been isolated to identify their role in the developing spike of wheat. Four genes were isolated from its gene family and designated as TaGAST1, 2, 3, 4. Genomic structure was analyzed to demonstrate chromosomal localization of TaGAST genes and evolutionary relationships were also verified with GAST genes in other plant species. RT-PCR was conducted to detect transcriptional changes of TaGAST genes on external phytohormone. Each of TaGAST genes showed considerable changes in transcription level after GA, ABA, PAC treatment, respectively. Through Yeast two-hybrid assay, one protein for TaGAST1, and four proteins for TaGAST2 was isolated as putative interactive proteins in wheat spikes just before and after emergence.

Brachypodium distachyon is a temperate annual grass that has a short life cycle, a small genome size, self fertility, and a small physical stature. The relationship with major cereal crop including wheat, Brachypodium is considered as a monocot model plant. Recently, the cell wall composition of Brachypodium is reported closely related with maize and Miscanthus giganteus. Therefore, Brachypodium is emrging as a powerful model plant for bioethanol production. Here, Brachypodium was chronically irradiated with the doses of 50 Gy, 100 Gy, 150 Gy, 200Gy, 250Gy, and 300 Gy. Plant height and fresh weight were observed dosage-dependent negative effect. However, tiller number and internode diameter were found to be increased their value as compared to control. The cell wall yield showed a decreased tendency with dosage-dependent negative, but cell wall yield of 50 Gy and 200 Gy were detected higher than control. The lignin content of irradiated Brachypodium stem was reduced with dosage incease The ratios of lignin content to control were 97.6% (50 Gy), 91.9 (100 Gy), 87.3% (150 Gy), 89.4% (200 Gy), 81.6% (250 Gy), 85.2% (300 Gy). SEM image analysis demonstrated that cell size of 300 Gy plant was decreased by 45% of control. RT-PCR was performed to analyze transcript accumulation of lignin pathway related genes with irradiated Brachypodium stem. CCR, PAL, C4H, and 4CL were detected at least 2 times higher expression than control at 150 Gy, 200 Gy, 250 Gy. The preteatment and enzyme hydrolysis will be discussed for bioethanol production.

The metalloid arsenic (As) and the hevy metal cadmium (Cd) are ubiquitously found at low concentrations in the earth, while high concentrations of the both elements in soil and crop are severe dangerous to human health. We have tried to retrieve RING E3 ligase gene, which is believed to regulate substrate proteins in As or Cd uptake via ubiquitin 26S proteasome pathway, related to inhibit metal ion transport system. A total of 48 rice RING E3 ligases were randomly selected and then conducted semi-quantitative RT-PCR for their expression patterns as exposed to As and Cd treatments. We discovered one gene, Oryza sativa heavy metal induced RING E3 ligase 1 (OsHIR1) that was significantly up-regulated against both treatments. A total of 31 positive interaction clones with OsHIR1 were screened depending on their strong α-galactosidase activity via yeast-two hybrid screen. Bimolecular fluorescence complementation analysis evidenced that the OsHIR1 protein was clearly interacted with each of six partner protein including aquaporin tonoplast intrinsic protein 4;1 (OsTIP4;1) in the plasma membrane. Protein degradation assay showed that OsHIR1 strongly degraded the protein level of OsTIP4;1 via ubiquitin 26S proteasome system. Heterogeneous overexpression of OsHIR1 in Arabidopsis showed As- and Cd-insensitive phenotype. In addition, the transgenic plant showed low levels of As and Cd accumulation than the control plant in leaf and root. Here, we report the novel finding that OsHIR1 E3 ligase positively regulates OsTIP4;1 related to As and Cd uptake.

The main objectives of IRRI’s variety development should meet the needs of customers/farmers from diverse rice sectors in each target region. The dynamic market change asks rapid variety development with highly valued QTLs/genes. Molecular breeding implemented through the efficient crossing, high throughput genotyping and rapid generation advancement will provide packages to breeders to develop new varieties quickly and more economically. The more efficient and cost-effective marker-assisted backcrossing service will provide the more opportunity for the success in molecular breeding platform. To make MABC system more successful, the development of molecular marker system for the high-throughput SNP genotyping is must. Currently Genotyping Service Lab (GSL) of IRRI provides high-throughput SNP genotyping service using BeadXpress and Fluidigm system. Meanwhile, the linked SNP markers for the specific traits are being developed. For abiotic stress tolerances, the markers for submergence, drought, heat, anaerobic germination, salinity, and phosphorus deficiency for Fluidigm system are being developed and tested in variety diversity panel and segregating populations. In MABC, due to the high number of crossings, the labor- and space-saving crossing system is being developed. As a result of an integrated MABC platform will speed up the development of pre-breeding line which are containing single or multiple QTLs/genes.

Rice (Oryza sativa) is an excellent model monocot with a known genome sequence for studying developmental seeds. In the study, the seeds of 10th day after flowering (DAF) were conducted RNA-Seq of the variety Shindongjin and Sugary mutant using RNA-seq technique. Approximately 202 and 214 million high-quality paired-end reads (101-bp in size) were generated in Shindongjin and Sugary mutant, respectively. Comprehensive analysis on the transcript levels of genes which encode starch-synthesis enzymes is fundamental for the assessment of the function of each enzyme and the regulatory mechanism of starch biosynthesis in seeds. Quantitative real-time PCR was also used to validate the expression profiles of 28 rice genes encoding six classes of enzymes, viz., ADPglucose pyrophosphorylase (AGPase), starch synthase, starch branching enzyme, starch debranching enzyme, starch phosphorylase, and disproportionating enzyme at different developmental grain- filling stages (DAF 1-14) between Shindongjin and Sugary mutant. The results showed that the expression of most of starch synthesis genes were up-regulated except the cytosolic AGPase small subunit2b (AGPS2b), which sharply decreased at grain-filling stages in Sugary mutant. These results will expand our understanding of the complex molecular and cellular events in rice grain-filling stages and provide a fundamental understanding of future studies on developmental endosperm in rice and other cereal crops.

Abiotic stress is the major limiting factor of forage crops growth and yields. The objective of this work was to study the stress tolerance and regeneration capability of transgenic forage crops carrying a MsHSP23 gene, encoding a alfalfa mitochondrial sHSP protein. The expression of the MsHSP23 gene was confirmed in bacteria, recombinant mHSP23 conferred tolerance to salinity and arsenic stress. Furthermore, mHSP23 was cloned in a plant expressing vector and transformed into forage crops such as alfalfa, tall fescue and bent grass. The transgenic plants exhibited enhanced tolerance to salinity and arsenic stress conditions. In comparison to wild type plants, transgenic plants were exhibited significantly lower electrolyte leakage. Moreover, the transgenic plants had superior germination rates when placed on medium containing arsenic. Taken together, these overexpression results imply that mHSP23 plays an important role in salinity and arsenic stress tolerance in transgenic forage crops. This approach could be useful to develop stress-tolerant plants including forage crops.

Legume and rhizobia symbiosis plays an important role in conversion of atmospheric dinitrogen to ammonia. On a global scale, thin interaction represent a key entry point for reduced nitrogen into the biosphere, and as a consequence this symbiosis in important in both natural and agricultural systems. Symbiotic development of nodule organ in triggered by chito-oligosaccharide signals(Nod factors) from the bacterium which are perceived by the legume root. Understanding the molecular and cellular processes that underlie Nod factor perception is one focus of legume biology. Although forward genetics has proved to be an important tool to elucidate key players in Nod factor perception, we still know relatively little regarding the functional networks of genes and proteins that connect the earliest steps of Nod factor perception to immediate downstream outcomes. To identify genes and proteins that link Nod factor perception to cellular and physiological responses we are taking a discovery-based strategy on large-scale transcriptome profiling using RNA sequencing in the roots of Medicago truncatula in response to Sinorhizobium meliloti. Functional characterization of a number of candidate genes is currently in progress to further examine their role in nodulation.