Rice is one of the most important food crops in the world, and has been used as model monocots for genetic studies, because of its relatively small genome size. We have previously reported the different functions of several RING (Really Interesting New Gene) proteins to respond the various abiotic stresses. In order to study a regulation of RING proteins in rice under ionizing irradiation such as gamma ray (GA), we have identified the expression patterns of these genes by RT-PCR. We found Gamma-ray induced RING finger protein (OsGRP) gene, which were associated with cytosol by subcellular localization analysis. in vitro ubiquitination assay revealed that OsGRP possess E3 ligase activity. Also, we demonstrate that C196A point mutation in the RING finger domain of OsGRP can have a critical effect to the breakdown of structural integrity in RING constructs. To identify the interaction partners for OsGRP in protein-protein interactions, we found the seven genes interacted with OsGRP by Yeast Two Hybrid method. To examine the GA-influence of interaction partners by RT-PCR, two genes were specifically down-regulated in rice during GA treatment. These interaction partners were identified the reliable interactions and subcellular localizations via BiFC method. Interestingly, five genes associated with plastid, while two down-regulated genes associated with cytosol and plastid. These results of OsGRP based on genetic approach might provide a clue to understanding the GA responsive mechanism in rice.

Direct seeding of rice is increasingly being practiced in both rainfed and irrigated areas because of labor shortage for transplanting and opportunities for crop intensification. However, slow seed germination and delayed seedling establishment will become a major problem for rice production in flood-prone lowland areas as sowing method shifts from transplanting to direct seeding. Identification of anoxia-induced ethylene response factors is suggestive because genes belonging to this gene family play a crucial role in rice tolerance to submergence. In this study, genetic structure variability of three AG related genes, Sub1 (Sub1A, Sub1B, Sub1C), Sub2 (OsGAPPH) and Ramy3D were examined by using whole-genome resequencing data of 84 accessions of rice core set. Some new SNPs and InDels found in exon part of anaerobic germination related genes in the present study would be useful in developing markers to identify the submergence resistant varieties in the future molecular breeding.

A male gametophyte, or pollen develops in the anther, and its development plays an important male reproductive process in flowering plants. A properly designed transgene construct can help to tailor transgene expression in plants by altering the expression strength, timing, and location. In this process, the promoter plays a pivotal role in controlling transgene expression. In this research, the promoter regions of rice anther/pollen-specific genes, named as OsMSP1 to OsMSP11,were selected from the microarray data sets covering 4 developmental stage of male gametophyte and then used for the construction of vector by Gateway cloning method and transformed into rice and Arabidopsis. All 11 promoters in rice and 9 in Arabidopsis were displayed as anther/pollen-specific/preferential genes by GUS assay and RT-PCR analysis. Three out of 11 promoters showed consistent results with published data. In this study, we demonstrated on eight new anther/pollen-specific or -preferential promoters (OsMSP1, OsMSP2, OsMSP3, OsMSP4, OsMSP5, OsMSP6, OsMSP8, and OsMSP9, which have not been reported before. Although the expression pattern of different genes active in pollen grains is diverse and complex, these experimental results would be helpful to understand the molecular mechanism of regulatory elements in rice microspore/pollen-specific genes.

In order to better understand the biological systems that are affected in response to cosmic ray, we conducted the weighted gene co-expression network analysis with module detection method. By using the Pearson’s correlation coefficient value, we were evaluated the complex gene-gene functional interactions between 680 CR-response probes from integrated microarray datasets, which included large-scale transcriptional profiling of 918 microarray samples. These probes were divided into 6 distinct modules that contained 20 enriched function such as oxidoreductase activity, response to stimulus and stress, and hydrolase activity. Especially, module 1 and 2 commonly showed the enriched annotation categories such as oxidoreductase activity, including the enriched cis-regulatory elements known as ROS specific regulator. These results suggest in module1 and 2 that ROS-mediated irradiation response pathway are affected by CR. We found the 243 irradiation-dependent probes, which were exhibited the similarities of differentially expressed patterns in various irradiation microarray datasets, and RT-PCR for confirmations of several irradiation-dependent genes were exhibited the similar expressed patterns in rice by CR, gamma ray and Ion beam treatments. Interestingly, these genes were differentially expressed by non-gravity. Moreover, we were identified the co-regulations between several irradiation-dependent genes and functional interacted genes in the CR-responsive network by various GA treatments such as different conditions of dose and treatment time. These results of network-based analysis might provide a clue to understanding the complex biological system of CR.

Alfalfa (Medicago sativa L.) is one of the most important forage crops in the world and it’s has been known as the best feed materials for dairy cows and other high valued animals. The new uses of alfalfa are being explored as bio-energy, food, medical and biochemical uses. R2R3-type MYB transcription factors play important roles in transcriptional regulation of anthocyanin biosynthesis. The R2R3-type IbMYB1 is known to be a key regulator of anthocyanin biosynthesis in the storage roots of sweetpotato. We previously showed that the expression of IbMYB1a led to anthocyanin pigmentation in tobacco and Arabidopsis. In this study, we generated transgenic alfalfa plants expressing the IbMYB1a gene under the control of CaMV 35S promoter. Overexpression of IbMYBa in transgenic alfalfa produced strong anthocyanin pigmentation in seedlings and generated a deep purple color in leaves, stems, roots, and even in seeds. High performance liquid chromatography (HPLC) analysis revealed that IbMYB1a expression led to the production of cyanidin as a major core molecule of anthocyanidins in alfalfa, as occurs in the purple leaves of sweetpotato (cv.Sinzami). We also examined expression of several structural genes in the anthocyanin biosynthetic pathway in alfalfa by RT-PCR analysis. In this presentation, we will further present molecular and biochemical characterization in IbMYB1a-overexpression lines. This result shows that the IbMYB1a transcription factor is sufficient to induce anthocyanin accumulation in the forage legume alfalfa plants.

Capsinoids which were found recently in non-pungent pepper show the same biological effects as capsaicinoid including anticancer and anti-obesity. A precursor of capsaicinoids, vanillyl alcohol, is known to be produced by mutations in the p-aminotransferase (pAMT) gene. In the previous study, we showed that capsinoid production is also controlled by the capsaicin synthase (CS) gene. However correlation between the CS gene expression and capsinoids contents has not been fully understood. This study was conducted to elucidate correlation between the expression level of CS gene and capsinoids contents. Through germplasm screening, we identified one C. chinese pepper cultivar, SNU11-001, which contained capsinoids as much as C. annuum ‘CH-19 Sweet’. SNU11-001 was crossed with five Capsicum cultivars (ECW, Takanotsume, Yuwolcho, Habanero and Jolokia) containing different levels of capsaicin, ‘ECW’ is non-pungent pepper line, and ‘Takanotsume’ and ‘Yuwolcho’ have mild pungency, and ‘Habanero’ and ‘Jolokia’ is known to be included in the most pungent pepper lines. When we analyzed the expression of CS and pAMT genes using the six Capsicum cultivars, the expression levels of CS were higher in pungent Capsicum cultivars. To test whether the expression levels of CS also control capsinoids contents, we will analyze several F2 populations derived from crosses between SNU11-001 and Capsicum cultivars containing different levels of capsaicin.

Rice is not only a model plant of monocots but also one of the most important crops all over the world. Despite the importance of leaf shape for achieving effective plant architecture for photosynthesis, little is known about the genetic mechanisms that determine leaf morphological characteristics. Explanation of the genetic basis of the control of leaf shape could be of use in the manipulation of crop traits, leading to increased crop production. Many mutants related to leaf morphology have been identified and classified according to their function in determining leaf morphology. search on the genetics of leaf development has used mutagensis to create loss-of-function mutations that change leaf shape. To understand the molecular mechanism of leaf morphogenesis, we identified a rice mutant gene, which was characterized by a phenotype of narrow leaves. While the mutation resulted in reduced leaf width, no significant morphological changes at the cellular level in leaves were observed, except in bulliform cells. The gene locus guess that it encodes a adenosine kinase, which displays sequence homology with ribokinase pfkB like superfamily. To test function of gene, we cloned gene which have 1140 nucleotides and 379 amino acids. This gene was transcribed in various tissues and was mainly expressed in panicles and leaves. NAL7, NAL1 and SLL1 were found to be downregulated, whereas OsAGO7 and NRL1 were upregulated in the mutant. These findings suggested that there might be a functional association between these genes in regulating leaf development.

Glycoalkaloids are a family of toxic secondary metabolites present in the plants of solanaceae family, which serve for plant defense. Two major glycoalkaloids present in plants are a-solanine and a-chaconine. The upper safe limit of glycoalkaloids for human consumption is 20mg/KG FW and its excess may cause severe health disorders. Light is the major factor known to increase the glycoalkaloid content in post harvest potato tuber. Glycoalkaloid pathway is not completely understood. Hence, identification and characterization of SGA biosynthetic genes and the genetic factors that control their expression levels assumes significance. Present investigation was focused on the study of expression pattern of key genes in steroidal glycoalkaloidal pathway under various light qualities in potato (Solanum tuberosum L). Two potato cultivars Atlantic and Haryeong which accumulates low and high glycoalkaloids respectively were used to check the levels of gene expression under various light qualities viz., red, blue, white, green, yellow, purple, UV light and in dark at different time intervals. Expression of three genes viz., SGT1, SGT2 and SGT3 which are directly involved and four other genes, HMG1, SQS1, SMT1 and SMT2 in the pathway envisaged to be indirectly involved in the glycoalkaloid formation was quantified by RT PCR. Varietal variation in the expression among the genes was observed in different light qualities. White, red and green light compared to other light qualities majorly contributed for the increased expression of genes for glycoalkaloid accumulation at different time intervals. Importantly, there is no significant transcript accumulation of these genes in dark condition. However, more efforts would be extended for further understanding of glycoalkaloid accumulation under light.

Rice is a staple food crop in the world. A number of agronomically important traits including enhancement of stress tolerance, quality improvement, and nutrition value increases have been introduced to rice. In this study, an Oryza sativa cDNA containing a U-box motif was cloned; its deduced amino acid sequence was compared to that of other U-box genes and indicated that encodes a U-box-containing E3 ligase. E3 ligases are structurally divided into three groups. We isolated the OsUPS gene from rice (Oryza sativa). The OsUPS protein has domain which is a single~70-amino acid region of the protein and GKL domain containing conserved Glycine, Lysine/ Araginine residues and leucine-rich feature. A full-length expression of OsUPS was up-regulated in the rice plant and in cell culture in the absence of phosphate. To express the OsUPS cDNA, it was inserted into the pGEX-2T vector. And the gene was expressed in E.coli strain BL21 (DE3). Induced after 3h of IPTG treatment and was isolated by affinity chromatography. Using the GUS reporter genes regulated by the OsUPS promoter, we have carried out the analysis of transcriptional and spatial regulation of gene expression. To investigate the function of these genes, the CaMV 35S promoter-driven these genes were introduced into Arabidopsis and rice via Agrobacterium tumefaciens-mediated gene transformation. We found that full-length expression of OsUPS was up-regulated in both rice plants and cell culture in the absence of inorganic phosphate (Pi). A self-ubiquitination assay indicated that the bacterially expressed OsUPS protein had E3 ligase activity, and subcellular localization results showed that OsUPS was located in the chloroplast. These results support the notion that OsUPS plays an important role in the Pi signaling pathway through the ubiquitin-26S proteasome system.

Somatic embryogenesis is a process where a plant or embryo is derived from a single somatic cell or group of somatic cells. Application of this process include: clonal propagation of genetically uniform plant material; elimination of viruses; provision if source tissue for genetic transformation; generation of whole plants from single cells called protoplasts; development of synthetic seed technology. In this study tissue culture was carried out for mass propagation of cassava using somatic embryogenesis. For tissue culture set up, we used cassava variety (“Rayong5”, “Rayong7”, “Rayong9”, “Rayong11”) developed in Rayong Field Crop Research Center(RYFCRC). In induction of callus step, the callus formed from each cassava variety. “Rayong 7“ showed the highest induction rate of 95%, while induction rate of other varies were ranged from 50% to 85%. In the case of weight of callus ”Rayong5“ has the highest weight. Results in the present study would be useful in mass propagation of cassava by somatic embryogenesis.

In order to adapt to various environmental stresses, plants have employed diverse regulatory mechanisms of gene expression. Epigenetic changes, such as DNA methylation and histone modifications play an important role in gene expression regulation under stress condition. It has been known that some of epigenetic modifications are stably inherited after mitotic and meiotic cell divisions, which is known as stress memory. To understand molecular mechanisms underlying stress memory mediated by epigenetic modifications, we developed Arabidopsis suspension-cultured cell lines adapted to high salt by stepwise increases in the NaCl concentration up to 120 mM. Adapted cell line to 120 mM NaCl, named A120, exhibited enhanced salt tolerance compared to unadapted control cells (A0). Moreover, the salt tolerance of A120 cell line was stably maintained even in the absence of added NaCl, indicating that the salt tolerance of A120 cell line was memorized even after the stress is relieved. By using salt adapted and stress memorized cell lines, we intend to analyze the changes of DNA methylation, histone modification, transcriptome, and proteome to understand molecular mechanisms underlying stress adaptation as well as stress memory in plants.

The utilization of several genetic resources is the most important for developing rice, such as high yield seeds or various stresses. We used an efficient system to create rice mutant by Ac/Ds transposon insertion mutagenesis, such as selected homozygous mutant in dwarf phenotypes. We reported here the identification of function of dwarf OsGASD gene (Oryzasativa Gibberellin Acid Sensitive Dwarf). OsGASD gene encodes a 344 amino acid polypeptide and nohomology proteins in GeneBank. The osgasd mutnat was sensitive to exogenous gibberellic acid (GA) level. We performed experiment to controlled expression the OsGASD gene, its role in plant development, a quantitative analysis of endogenous GA content and sensitivity to GA. The osgasd mutant includes smaller amount of active GAs than wild-type. osgasd mutant plant of GA biosynthesis pathway causes GA deficiency and dwarf plants ,and endogenous GA suppliance can restore the wildtype phenotype in this mutant. The result indicated that OsGASD gene regulated the elongation of shoot, stem and plant height. The increased expression of OsGASD gene dramatically induces expression of the factors associated with GA biosynthesis such as CPS, KO, KAO, GA20ox and GA2ox, whereas osgasd mutant suppression of the factors associated with GA biosynthesis, loading to dwarf phenotypes. That applied GA3 at the plant development stage to survey the response of OsGASD gene to GA3. We suggest that OsGASD gene is related to factors of GA biosynthesis pathway regulating rice internodes development.

Arabidopsis atDjC53 and atDjC32 gene DnaJ-like protein homologous to DnaJ-like protein was characterized for the functional analysis of DnaJ-like protein. It was shown that atDjC53 and atDjC32 RNA expression is induced by heat shock stress and atDjC53- and atDjC32-GFP was targeted to the nucleus of protoplasts. The atDjC53 and atDjC32 promoter (1 kb) was isolated and fused to the GUS reporter gene to investigate gene regulation of atDjC53 and atDjC32 specific to heat shock stress or to developmental organ in the transgenic lines. RNAi and overexpression construct was employed to generate atDjC53 and atDjC32 knock-out plants for the study of their function. Molecular function of atDjC53 and atDjC32 is discussed in relation to heat shock and also developmental stages in Arabidopsis.

We used a microarray dataset that is deposited in the public database to evaluate plant responses to heat stress and selected two genes, OsSHSP1 (Os03g16030) and OsSHSP2 (Os01g04380), that are highly expressed under heat stress in rice. OsSHSP1 and OsSHSP2 gene transcripts were highly induced in response to salt and drought. In addition, OsSHSP1 and OsSHSP2 gene transcripts were induced under ABA and SA. Subcellular localization of proteins of 35S::OsSHSP1 were associated with the cytosol, whereas those of and 35S::OsSHSP2 were associated with the cytosol and nucleus. Heterogeneous overexpression of both genes exhibited higher germination rates than those of wild-type plants under the salt treatment, but not under heat or drought stress. The network of both genes harboring 9 sHSPs as well as at least 13 other chaperone genes might support the idea of a role for sHSPs in the chaperone network. Our findings might provide clues to shed light on the molecular functions of OsSHSP1 and OsSHSP2 in response to abiotic stresses, especially heat stress.

Off-type rice plants occurring in farm fields cause yield loss due to competition with cultivated rice, in addition to hindering field management and harvest work. This study aimed to observe the agronomic characteristics and trace the origins of off-type rice plants using molecular markers. A total of 116 rice accessions, comprising 35 off-type plants collected from Korean farm fields, 19 Korean commercial cultivars, 12 Korean land races, and 50 weedy rice collections, were phenotyped and genotyped using selected SSR and Subspecies Specific (SS)-STS markers. The results showed that the plant height, culm length, and leaf length of off-type rice plants were larger than those of cultivated rice, which is the typical phenotype of weedy rice. However, off-type plants were highly sterile, as opposed to weedy rice, which were highly fertile. Genotype analysis with SSR and SS-STS markers revealed that off-type rice plants were heterozygous at most of the tested marker loci, suggesting that the off-type rice plants may have originated from natural outcrossing. The genotypes of off-type rice plants were closely related to both weedy and cultivated rice, and the phylogenetic analysis revealed that the relationship of the clustered group of offtype rice plants is intermediate between Indica type weedy rice and Japonica type commercial varieties. These results suggested that off-type rice plants collected in Korean farm fields might have originated from natural outcrossing between Indica type weedy rice and the cultivated Japonica type commercial varieties.

Low temperature is a major factor restrict to growth and limiting productivity of rice crops. We used a cDNA microarray approach to monitor the expression profile of rice (Oryza sativa) under chilling stress and identified 20 chilling inducible genes in previously study. Ten such genes encoding bHLH, metal transporter and, zinc finger protein with unknown functions showed a significant change in expression under various abiotic stresses. Among them, OsCHI1 (Os07g15460), OsCHI2 (Os02g43660), and OsCHI3 (Os01g61160), were selected for further study. They have structural features such as metal-binding signature sequences in their protein sequences, and OsCHI genes were expressed in root of rice seedling and induced in chilling and salt or drought. Expression of OsCHI1, OsCHI3 and OsCHI2 were targeted to membrane and ER when transiently expressed in tobacco cell, respectively. The Arabidopsis (Arabidopsis thaliana) transgenic plants overexpressing showed increased tolerance to salt and drought stress in the seed germination and root elongation than that of wild type. This comprehensive study provides insight into the biological function of OsCHIs, which may be useful in understanding how rice plants adapt to unfavorable environmental conditions.

60년대와 70년대의 녹색혁명은 전통적인 육종의 성과이지만, 최근의 육종기술은 유용유전자의 유전자형을 활용한 분자표지 마커를 활용하고 있다. 목적 형질을 갖고 있는 계통을 선발하는 MAS(Marker assisted selection) 마커는 많은 육종가들에 의해 활용되고 있지만, 유전자의 SNP(Single nucleotide polymorphism)을 활용한 MAB(Marker assisted backcross) 마커는 거의 활용되고 있지 않다. SNP 마커는 단 하나의 염기서열의 차이를 구별할 수 있어, 유전적으로 매우 가까운 계통들도 구분할 수 있으며, 자동화 분석이 가능하다는 점에서 활용도가 높다. 솔젠트(주)에서는 내수 및 종자수출에서 차지하는 산업적 비중이 가장 큰 채소작물 중 고추 병저항성 관련 유전자를 활용한 multiplex 고추병 진단 제품을 개발했다. 이 제품에는 오이모자이크바이러스(CMV), 담배 etch 바이러스(TEV), 토마토반점위조바이러스(TSWV), 토바모바이러스(TMV), 세균성점무늬병(BS), Chilli venial mottle virus(ChiVMV), 고추역병에 관련된 저항성 유전자의 SNP을 활용해 개발했으며, 본 제품을 활용해 고추 육종가들이 쉽고 간편하게 육종 소재에 적용해 신품종 육종에 도움을 줄 것으로 기대한다.

Rice hulls remain closed throughout the ripening period to maintain internal humidity of the grains. An Open-hull sterile mutant was induced by N-methyl-N-nitrosourea(MNU) treatment on Sinsunchalbyeo rice, a japonica type. This mutant showed open hulls even in the ripening stages and fully mature grains. In addition, several altered characteristics were observed, including of narrowed palea, decreased grain size, partial pollen sterility and erect panicle. Microscopic analysis showed that the palea was positioned slightly inside the lemma, and the size of palea decreased in the mutant. Genetic analysis of F2 and F3 segregation populations derived from the cross between the Open-hull sterile mutant (Oryza sativa ssp. japonica) and Milyang23 (O. sativa ssp. indica) indicated that the Open-hull trait was controlled by a single recessive allele. The fine-mapping with STS (sequence tagged site) markers revealed that the mutant gene was located on the short arm of chromosome 3. We were able to narrow it down until 30.6Kb where three candidate genes were found.

Plant growth under water-deficit conditions adversely affects many key processes. Efforts to understand drought stress-related defense mechanisms have revealed a host of plant genes using molecular approaches in rice. Here, we report the novel finding that OsCTR1 E3 ligase regulates both chloroplast-localized chloroplast protein 12 (OsCP12) and ribosomal protein 1 (OsRP1) in protein levels and subcellular localization. The results of a yeast-two hybrid assay, bimolecular fluorescence complementation assay, ubiquitination assay, subcellular localization, and a protein degradation assay support the hypothesis that OsCTR1 functions in trafficking inhibition and proteolysis of OsCP12 and OsRP1 via the ubiquitin 26S proteasome pathway. Heterogeneous overexpression of OsCTR1 in Arabidopsis showed ABA-hypersensitive phenotype in seed germination, seedling growth, and stomatal closure. The transgenic plants also exhibited improvement of water-deficit tolerance with an accumulation of hydrogen peroxide production. These results demonstrate that the OsCTR1 E3 ligase might positively regulate the cellular functions of OsCP12 and RP1 related to photosynthesis under drought stress conditions in rice.

Hybrid sterility is a critical barrier of inter-subspecific crosses in rice. However, hybrid sterility can be overcome by wide compatibility variety. The HWC-line of rice had slender grain shape, tall culm length, wide compatibility with both indica and japonica cultivars. For QTL analysis of HWC-line, two F2 populations were derived from the crosses between the HWC-line and each of two Korean variety, Dasan (Korean Tongil-type cultivar) and Hwacheong (temperate japonica cultivar). In the cross between HWC-line/Dasan (HD), 93 STS markers and 13 SSR markers were mapped on 12 chromosomes. In the population from HWC-line/Hwacheong (HH) cross, 28 STS markers, 29 SSR markers and 1 FNP marker were mapped on 11 chromosomes. Eight agronomic characters were evaluated for QTL analysis in two F2 populations and parents. The F2 population from HD cross revealed 21 M-QTLs and 3 E-QTL for culm length, spikelet per panicle, spikelet fertility, grain length, grain width, grain shape and 100 grains weight. 8 QTLs of culm length, grain length, grain width and grain shape were newly detected in this study. In the F2 population from HH cross, 17 M-QTLs were detected for culm length, panicle length, spikelet fertility, grain length, grain width, grain shape and 100 grains weight. 6 QTLs of culm length, grain length, grain width and grain shape were newly found in this study. These QTLs will be able to provide basic information on putative functional genes related with agronomic characters and promote breeding new rice cultivar. HWC-line could be used as bridge for inter-subspecies crosses and in hybrid breeding.