8th World Congress on Plant Genomics and Plant Science August 10-11, 2018 Osaka, Japan

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Lerma S J Maldia has expertise in tree population genetics and molecular phylogenetics and phylogeography. Through support funds from various government agencies and the university, she has recently established the Forest Genomics Laboratory of the College of Forestry and Natural Resources of UPLB, in support to the vision of competitive research and excellent instruction of the university. Currently, she is leading a number of similar genetic studies on various indigenous high value forest trees for their genetic conservation and sustainable utilization.

Abstract:

Statement of the Problem: Oaks are ecologically important components of broadleaved temperate forests of Japan. They are common planting species for forest restoration, greening programs and establishment of forest parks. However, genetic risk involved in planting without knowledge of the origins of plant materials has been a concern over these activities. Studies showed that introduction of foreign genotypes into new environment during restoration could result to loss of local adaptation, mortality due to maladaptation, genetic pollution or pose threats to regional biodiversity.

Methodology & Theoretical Orientation: To offset these risks, establishing transfer (or seed) zones or regions within which planting stocks can be moved with least or no negative effects on the average fitness of population must be carefully considered. Seed zoning have generally used edaphic and climatic data, ecophysiological data, and/or results of provenance tests. Recently, the utility of geographic patterns of genetic variations revealed by molecular markers for zoning in restoration has been recognized. In this study we determined the genetic variations at nuclear and chloroplast genomes of native white oaks species (Genus Quercus, Section Prinus; Q. aliena, Q. crispula, and Q. serrata), including some of their variants in Japan, in order to recommend genetic guidelines for transfer of planting materials for these oaks species for restoration.

Findings: The genetic structure and distribution of chloroplast haplotypes of the studied oaks species displayed generally distinct geographic genetic structures of northern and southern populations, following the north-south lineages of other deciduous broadleaved species in Japan, but with generally limited within-population diversity in northern populations. On one hand, the signals of genetic structure of nuclear genome seemed related to their range-wide distributions. The northsouth differentiation at nuclear genome was very weak for the widely distributed Q. serrata but very strong for the sparsely distributed Q. aliena and relatively fragmented in the south Q. crispula.

Conclusion & Significance: The sharp north-south difference in genetic backgrounds at two genomes suggests completely differentiated gene pools for these two large genetic populations, and thus, can be considered genetically distinct transferrable zones for planting stocks of Q. aliena and Q. crispula. Plant materials for restoration, therefore, should be restrictively sourced from and transplanted within each transfer zone and inter-planting between zones must be avoided.

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Cenek Novotny is an Associate Professor, has his expertise in the fields of physiology and biochemistry of bacteria, yeasts, ligninolytic basidiomycetes and bacterial and fungal phytopathogens. His research included the investigation of biofilms of bacteria and fungi, microbial interactions in mixed biofilms, antagonisms between microorganisms and biodegradation of recalcitrant organopollutants in water and soil. He is currently working at the Institute of Microbiology, CAS, Prague and the University of Ostrava, Czech Republic. He has coordinated five Czech and 14 international research projects and published 68 papers.

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Biocontrol agents represent an alternative to chemicals in the management of fungal crop diseases. A wide range of microorganisms can be used as biocontrol agents, mainly bacterial and fungal isolates and the biological control results from many types of interactions between organisms. In our study, Fusarium oxysporum f.sp. conglutinans was confronted with three different biocontrol agents, Trichoderma harzianum, Bacillus amyloliquefaciens and Pseudomonas aeruginosa in dual cultures. Bioassays and metabolites produced in the microbial interactions were screened by a Matrix-Assisted, Laser Desorption/Ionization (MALDI) mass spectrometry. T. harzianum exhibited the strongest inhibition of mycelial growth of F. oxysporum by overgrowing the pathogen in the later stages of co-cultivation. The metabolite profiles obtained in the case of T. harzianum and B. amyloliquefaciens were the result of an attack on the F. oxysporum mycelium by the antagonists by means of membrane-attacking peptaibols and a number of antimicrobial lipopeptides and siderophores, respectively. The biocontrol activity of T. harzianum and B. amyloliquefaciens consisted in their ability to suppress the production of mycotoxin beauvericin by F. oxysporum. In the case of P. aeruginosa, siderophores pyoverdine E/D and two rhamnolipids were produced as major bacterial metabolites. Under the conditions of a co-culture with F. oxysporum the production of rhamnolipides by the bacterium was blocked by the action of the fungal phytopathogen. The biocontrol of F. oxysporum by P. aeruginosa was weaker that those by T. harzianum and B. amyloliquefaciens. 

Biography:

Cheol Soo Kim is a Faculty Member in the Department of Plant Biotechnology at the Chonnam National University of South Korea. He focuses on researching the molecular genetic mechanisms underlying plant responses to adverse environments such as osmotic stress, drought and biotic stress. His work has led to the identification of genes for modifying the responses of crops to environmental stresses, which will ultimately lead to major contributions to agriculture and the environment.

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Proline (Pro) metabolism is important for environmental responses, plant development and growth. However, the mechanism of Pro in abiotic and biotic stress processes is unclear. Using the atrzf1 (Arabidopsis thaliana RING zinc finger 1) mutant as a parental line for T-DNA tagging mutagenesis, we identified a suppressor mutant, designated proline content alterative 13 (pca13) that suppressed the insensitivity of atrzf1 to abiotic stresses during early seedling growth. Pro content of pca13 was lower than in atrzf1, while the complementary lines were less sensitive to Abscisic Acid (ABA) and abiotic stresses compared to WT. Through the TAIL-PCR of pca13, it was shown that T-DNA inserted at site chromosome 2 which encodes cell wall enzyme. Under condition of biotic stress, pathogen resistance was significantly higher in pca13 compared to WT and atrzf1. Moreover, pca13 mutant was significantly higher in several important drought parameters including malondialdehyde level, ion leakage and water loss. The fluorescence signal of the green fluorescent protein (GFP)-tagged PCA13 was quite strong in the cell wall of the root cells of the transgenic seedlings. Additionally, the PCA13 promoter-β-glucuronidase (GUS) construct revealed substantial gene expression in the root tissues and apical meristem. Collectively, these findings prove that pca13 acts as a suppressor mutant of atrzf1 in the abiotic and biotic stress responses through the Pro metabolism. 

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Clustered regularly interspaced palindromic repeats (CRISPRs)/CRISPR associated (Cas) is currently known as a very powerful genome editing technology, and it is also success in many plants. Here, we first made Cas9 overexpressing tobacco plants through agrobacterium-mediated transformation method. Tobacco Cas9 overexpressing plants confirmed the Cas9 expression level by RT-PCR analysis. PDS (positive control) and eIF4E genes infiltrated in tobacco plants systemically. Expression of PDS and eIF4E confirmed the time course (0, 6, 24, 48, 72 hrs) by RT-PCR analysis. We are produced the new tobacco plants using transient expressing leaf through tissue culture, and the seeds are harvested and then selected from the selective germination medium (with antibiotic). If these results are successful, the TRV (Tobacco Rattle Virus) delivery system will be able to be very efficiently in many plant species. 

Biography:

Hye-Eun Lee is conducting the research on the development of molecular marker in various vegetables. She works at the National Horticultural Science Institute in South Korea, and now She studies about analysis of gene expression using Chinese cabbage and construction of genetic population and linkage genetic map related to agronomic traits in onion

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Ethylene responsive element binding protein (EREBP) is a major group of the AP2/ERF family and plays significant roles in the regulation of abiotic and biotic stress responses. StEREBP is cloned from potato (Solanum tuberosum L.) and characterized to response in abiotic stress and hormone regulation. In this research, we constructed Chinese cabbage StEREBP overexpressing plants using the Agrobacterium-mediated  transformation. Six transgenic plants are confirmed the T-DNA insertion by Southern blot analysis and RT-PCR analysis. Phenotype of transgenic plants is not different compared with control plant. The reverse transcription PCR (RT-PCR) showed that cold stress and ABA related genes are increased but ethylene related genes are decreased in StEREBP overexpressing Chinese cabbage plants. The StEREBP responded to abiotic factors and hormones suggested that they possibly had diverse roles in stress and hormone regulation of Chinese cabbage.

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Rimi Yamaguchi has completed both Master and Bachelor of Bioresource and Bioenvironmental Sciences in 2017 and in 2015, respectively from Kyushu University, Japan.

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Pine Wilt Disease (PWD) caused by the Pine Wood Nematode (PWN) Bursaphelenchus xylophilus, is one of the most serious forest disease in East Asia and Europe. Japanese black pine Pinus thunbergii is highly susceptible to PWD and therefore resistant trees have been selected to cope with this disease. Several studies have assessed the histological responses or transcriptional changes in resistant and susceptible trees. However, the underlying mechanism of resistance of pine is still not fully understood. The purpose of this study was to identify mechanisms involved in the resistance of pine from a perspective of the host-pathogen relationships. Here, we identified multiplication of PWN in two resistant and one susceptible tree varieties inoculated PWN and then performed the dual RNA-sequencing (RNA-seq) analysis. The susceptible trees showed wilting of leaves at 7 days post inoculations (dpi), and the number of PWN in trees was higher in the susceptible trees than the resistant trees at 14 dpi. RNA-seq performed on six cDNA libraries from two resistant trees and one susceptible tree at 1 and 3 dpi. A total number of 277,503,322 read pairs obtained from all cDNA libraries were mapped to the PWN genome. The percentages of reads mapped to PWN genome ranged from 0.11 to 0.36%, which indicated that we could reveal gene expressions of both hosts and pathogens simultaneously. Their expression profiles suggest that there is a difference in mechanism of resistance between resistant tree varieties against infection of PWN.

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Van Tinh Nguyen is currently pursuing PhD in Department of Applied Biology of Chonnam National University, South Korea. His major is plant genomics functional researches which involved in abiotic stress.

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Cysteine (Cys) and AMP are products of sulfate metabolism. Cys is an essential amino acid for protein and peptide synthesis or reduced sulfur donor for biosynthesis of methionine, coenzyme or cofactor, while AMP (Adenosine Monophosphate) is a monomer in the production of RNA and joins in many metabolism processes. However, whether theirs metabolism involved in abiotic stress adaptation in Arabidopsis remains largely unclear. Here, we identified atsmr17 (Arabidopsis thaliana sulfate metabolism related 17), a double mutant, that suppressed the insensitively of the parental line atrzf1 (Arabidopsis thaliana ring zinc finger 1) to glucose (Glc) treatment via reducing Cys and AMP accumulations. Under high Glc condition, the level of atsmr17 was significant induced. Losing function of atsmr17 leads to decrease the physiological phenotype including the germination rate, cotyledon greening, root and shoot differentiation. Interestingly, Glc response differently exhibited between root and shoot depend on atsmr17 levels. Besides, qPCR analysis of the genes involved in primary sulfate pathway exhibited significantly lower in atsmr17 leads to reduced 60% and 80% Cys contents compare to WT and atrzf1, respectively. Moreover, the atsmr17-overexpressing line displayed hyper-insensitive under high Glc concentration treatment manifested by the stress insensitive parameters and also by increased the proline, Cys, and AMP contents. Noticeably, apply exogenous Cys and AMP lead to rescue the phenotype of atsmr17 under high Glc treatment. Taken together, our results indicate that atSMR17 plays a role in high Glc response through modulating the sulfate metabolism in which related to Cys and AMP accumulations in Arabidopsis.

Biography:

Ji-Hee Min is currently pursuing her PhD in Applied Biology at Chonnam National University, South Korea under the direction of Professor Cheol Soo Kim. She is interested in plant stress responses via ubiquitination system which determine protein stability or degradation. Her research was concentrated upon defining the role of interactor of E3 ubiquitin ligase Arabidopsis thaliana RING zinc finger 1 (AtRZF1) in the abiotic stress response.

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The E3 ligase AtRZF1 is a component of drought-induced proline biosynthesis and drought sensitivity in Arabidopsis. In this study, we found that a novel AtRZF1-Coupling Protein 1 (AtRCP1) interacting with AtRZF1 by using Yeast two-hybrid and GST-pull down assays. Expression of AtRCP1 was strongly induced by Abscisic Acid (ABA), osmotic stress treatments. The green fluorescent protein-AtRCP1 fusion protein is localized in nucleus. Also, AtRCP1 has transcriptional activation activity and contains DNA binding domain indicating this functions as a transcription factor. The atrcp1 RNAi mutants were less sensitive to osmotic stress than the wild-type, whereas transgenic plants overexpressing AtRCP1 were hypersensitive during early seedling development, indicating AtRCP1 negatively regulates drought-mediated signaling. The expression levels of the stress markers or proline metabolism genes were altered both in AtRCP1 over expressing and knockdown plants. Biochemical study through ubiquitin binding assay and ubiquitination assay implied that UBA domain in AtRCP1 is required for binding of Ub chains and increase of the amount of ubiquitinated AtRZF1. Moreover, genetic studies showed that the AtRZF1 gene could rescue the ABA- and osmotic-insensitive phenotype of atrcp1 RNAi mutant suggesting E3 ubiquitin ligase AtRZF1 is epistatic to AtRCP1 in osmotic stress signaling. Our findings provide new insight into the mechanism by which water deficit controls proline accumulation and drought response in Arabidopsis.

Biography:

Ga Yun Jeong is fourth year student in Department of Applied biology, Chonnam National University of South Korea. She is studying in plant biotechnology research at the Cheol Soo Kim’s lab. She is interested in ABA and dehydration stress in Arabidopsis.

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The phytohormone Abscisic Acid (ABA) is the major component of abiotic stress tolerance in plants and coordinates a complex regulatory network enabling plants to cope with the stress. The precise roles of the homeodomain leucine zipper family of transcription regulator in plant stress are largely unclear. Here, we characterize the biological function of the AtHLZ1 Arabidopsis thaliana homeodomain leucine zipper-type: In ABA and dehydration responses. The expression of AtHLZ1 was strongly induced by ABA and osmotic stress. AtHLZ1 RNAi lines resulted in increased sensitivity to ABA and dehydration stress during seed germination and the cotyledon greening process. In contrast, The AtHLZ1-overexpressing transgenic plants were less sensitive to ABA and osmotic stress compared to Wild Type (WT). Interestingly, in the presence of ABA, the transcript levels of ABA receptor Pyrabactin Resistance-like 4 (PYL4) and PYL7 genes were enhanced reduction in AtHLZ1- overexpressing transgenic plants rather than in the WT and AtHLZ1 mutant. Thus, AtHLZ1 is involved in ABA and osmotic stress response through the ABA-dependent signaling pathway. The data show that AtHLZ1 is an important regulator in response to both ABA and dehydration stress in Arabidopsis.

Biography:

Ji Hyeon Min is an Assistant Researcher in Cheol Soo Kim’s laboratory at the Chonnam National University in South Korea. He focuses on researching crop development via CRISPR-Cas9 system to increase drought stress tolerance.

Abstract:

Watermelon is a major fruit vegetable around the world. Drought is an abiotic stress factor that affects the productivity and growth of crop plants. To enhance the tolerance of watermelon to drought stress, it is important to isolate stress tolerance-related genes. Recently, we characterized the gene for ubiquitin E3 ligase protein named Lagenaria siceraria RING Zinc Finger 1 (LsRZF1). In Arabidopsis, LsRZF1 is involved in the drought response through the proline metabolism-mediated pathway. In this study, we identified and characterized a watermelon (Citrullus lanatus cv. Gongdae) homolog of LsRZF1, designated GdRZF1. LsRZF1 antisense (lsrzf1) transgenic watermelon lines showed decreased GdRZF1 expression and were in sensitive to drought stress than the wild type. Decreased expression of GdRZF1 was also significantly influential in changes in drought-sensitive parameters including relative water content, ion leakage, chlorophyll content, malondialdehyde levels, proline content and the expression of drought stress responsive genes. Additionally, grafted plants showed more health leaves and larger leaf area as compared with the control during drought condition. These findings results that GdRZF1 is important for water deficit tolerance in watermelon. This is the report of the development by genetic engineering of watermelon tolerance to drought response.

Biography:

Jung In Kim is fourth year student in Department of Applied biology, Chonnam National University of South Korea. She is studying in plant biotechnology researching at the Cheol Soo Kim’s lab. She is interested in ethanol fermentation on anaerobic condition, also her work is related to post-translational modification stage

Abstract:

Ubiquitination is an important process related to the activities of enzymes which occurs in the post-translational modification stage. This process requires for the regulation of divers cellular functional through biological events including the response to hormone, apoptosis and abiotic stress. In this study, we characterized the biology functional of  AtEUL(Arabidopsis thaliana E3 Ubiquitin Ligase 1) in ethanol response. The initial result showed that, the mRNA level of AtEUL1 was dramatically induced by ethanol exogenous application. The transgenic plants overexpressing AtEUL1 were hypersensitive to ethanol response than wild-type during dark-grown seedling growth, whereas the ateul1 mutant was hypersensitive, indicating that AtEUL1 negatively regulates ethanol-mediated response of early dark-grown seedling growth. Additionally, the ateul1 mutant was significantly influential in ethanol sensitive parameters including hypocotyl length and the expression of ethanol metabolic genes. The taken together, the result suggests that AtEUL1, a ligase E3 enzyme, plays a function as ethanol regulator response in early seedling dark-grown of Arabidopsis development. 

Biography:

Do-Sun Kim has her expertise in plant transformation and tissue culture in introducing useful traits. She is working at National Institute Horticultural and Herbal Science in South Korea. Also, she is In-Charge of Chief of Vegetable Molecular Breeding Laboratory at the Institute and is active as expert Member at Korean Society for Plant Biotechnology.

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The edible vaccine derived from crop is easy to achieve safety and its price is cheap because of the mass production by agriculture system. To acquire edible vaccine fusion protein for colorectal cancer which is a serious disease in worldwide, we used various form of immunoglobulin Fc (IgM and IgA) and J chain. The J chain is expected to induce a fusion protein complex when co-expressed with other immunoglobulins. The Chinese cabbage (Brassica rapa), which is a vegetable, oilseed and fodder crop, grown especially in Asia and Europe, was selected to develop  edible vaccine due to the high protein  expression rate. The Agrobacterium transformation method was used to express each protein in the Chinese cabbage. The T0 transformants were acquired successfully. The three transgene (J-chain, EpCAM-IgM and EpCAM-IgA) was confirmed by PCR using gene specific primers. The transformation efficiency was approximately 3%. The crude proteins from T0 plants were extracted and the overexpressed proteins were detected by Western blot analysis. The over-expression of EpCAM-IgM and IgA were confirmed. T1 plants were also selected by hygromycin and its transgene expression was confirmed by immunoblot assay. We tried to develop vaccine fusion protein complex in Chinese cabbage by crossing T1 plants expressing J chain and immunoglobulin protein each. Chinese cabbage T1 plants each expressing EpCAM-IgM and IgA were crossed with the T1 plants expressing J chain. The F1 hybrid plants were acquired and its fusion proteins were confirmed. The fusion proteins were extracted and their expression levels in Chinese cabbage were estimated. The plants generated by this procedure will provide an efficient edible vaccine tool to prevent cancer disease in the near future. 

Biography:

Umme Qulsum is a Doctoral candidate in School of Materials Science, Japan Advanced Institute of Science and Technology. She is also affiliated to Department of Botany, University of Rajshahi, Bangladesh as an Assistant Professor. Her main interests include research on alternative splicing of RNA editing related family proteins and their effect on RNA editing in plants. She has experimental knowledge in plant molecular biology.

Abstract:

Arabidopsis is the most useful model plants in molecular biology. RNA editing is a post-transcriptional modification of genes that commonly occur in plant plastids and mitochondria. Alternative splicing is a post and co-transcriptional regulation of gene expression. In recent studies, it is found that the PPR family proteins are involved in RNA editing in plants. In flowering plants, not only PPR but also non-PPR proteins like MORF, ORRM and OZ participate in multiple RNA editing events. The PPR–RNA complex is organized into the editosome with several additional non-PPR protein factors. The aim of this study is to find out the frequency and extends of tissue-specific RNA editing events and expression of alternative splicing of these genes and their effect on protein structure and functionality. We collected samples of different tissues of different developmental stages of Arabidopsis. Gene expression analysis and sequencing were performed. I-TASSER was used for protein modulation study. We found editing events in eleven genes out of twelve of PPR family genes and only one genes out of nine of ZNF family genes. We found nine types of RNA editing events these events included possible intra-base transitions: C-to-U, U-to-C, A-to-I, A-to-C, A-to-U, G-to-A, G-to-C, U-to-A and U-to-G in targeted genes. Most of the editing events in seedling and leaf and less in stem tissues. In our study, most of the alternative splicing events were found in seedling and leaf. We also detected seven unannotated and new alternatively spliced isoforms among these five PPR and two ZNF genes that were confirmed by PCR and sequencing. RNA editing machinery may play important role in proteins diversity and functionality thus ultimately affecting plant physiology. This study suggests that tissue-specific expression of different alternatively spliced transcript happen even in different developmental stages. 

Biography:

Subhankar Bera is pursuing his PhD degree on plant science in Osaka Prefecture University. His PhD work is based on small RNA analyses trace the movement of small RNAs. He is building his experience in research and educational institution. He want's to devote and dedicate himself in dynamic and creative academic interest and to utilize his logical sense, analytical power, potentiality and skills those provide steady growth.

Abstract:

Parasitic plants are grown on hosts through haustorial formation to uptake water and nutrients for their survival and growth. During host-parasitic interaction plant endogenous mRNA and proteins also moved bi-directionally through parasitic interface tissue. Recent studies shown that parasitic plant accumulate miRNA in interphase tissue of host-parasitic complex to control/or regulate host gene through secondary siRNA production. However, there is no direct evidence of host-derived small RNAs moved to long distance tissue of parasite. The purpose of this study is to find out commonly shared long-distance moved small RNAs in both host and parasitic plants which may have functional regulation during and after host-parasitic interaction. We design experiments on (Cuscuta japonica-Glycine max) and (Cuscuta campestris-Arabidopsis thaliana) as different species of parasitic plants grow on different family of host plants. Small RNA-seq analyses and comparison of non-parasitic and parasitic tissue of both parasite and host plants showed that several small RNAs are produced from common gene family of hosts moved to long distance tissue of parasitic plant. Similarly, there are some orthologous genes of different parasitic plant species produced small RNAs from and moved to long distance tissues of hosts. Small RNA candidates target common of orthologous hosts genes also. By stem-loop PCR followed by Sanger sequencing validate small RNAs. Long distance movement was proved by cross-species detection of sRNAs. These results suggest that small RNAs are moved in bidirectional manner to control trans-species gene regulation.