Plant biotechnology including plant tissue culture is the most important science to rescue human beings from the crisis of biosphere of the earth which they will face in the 21st century. Global area other than Asia of trangenic crops increased more than 16 times from 1996 to 1998. In Asian countries, micropropagation using tissue culture techniques has been well developed and has contributed to agricultural production. Although researches on levels of laboratory and test field trial of transgenic crops have been performed actively in some Asian countries, areas of growing transgenic crops are still small in Asia countries except in China. In this paper, the status of plant biotechnology in global and Asian countries are reviewed and the future prospect of plant biotechnology in Asia and roles of Korean and Japanese plant scientists in it are discussed.

In order to evlauate feasibility of the gene tagging by the maize transposable element Ac in heterologous plant systems, we have investigated physical distances and directions of transposition of the element in Arabidopsis thaliana and tobacco cultured cell line BY-2. We prepared a T-DNA construct that carried a non-autonomous derivative of Ac with a site for cleavage by endonuclease I-Scel (designated dAc-I-RS element). Another cleavage site was also introduced into the T-DNA region outside dAc-I-RS. A number of transgenic Arabidopsis plants were generated, each of which had a single copy of the T-DNA at a different chromosomal location. To examine the pattern of transposition, three out of these transgenic plants were crossed with the Arabidopsis plant that carried the gene for Ac transposase and progeny in which dAc-I-RS had been transposed were isolated. After digestion of the genomic DNA of these progeny with I-SceI, sizes of segment of DNA were determined byd pulse-field gel electrophoresis. We also performed linkage analysis for the transposed elements and sites of mutations near the elements. Our results with three transgenic lines showed that 50% of all transposition events had occurred within 1,700 kilo-base pairs (kb) on the same chromosome, with 35% within 200 kb, and that the elements transposed in both directions on the chromosome with roughly equal probability. The data thus indicate that the Ac-Ds system is most useful for tagging of genes that are present within 200 kb of the chromosomal site of Ac in Arabidopsis. In addition, determination of the precise localization of the transposed dAc-I-RS element should definitely assist in map-based cloning of genes around insertion sites. In the present paper, we report typical examples of such gene isolation studies.

In rice, limited efforts have been made to identify genes by the use of insertional mutagens, especially heterologous transposons such as the maize Ac/Ds. We constructed Ac and gene trap Ds vectors and introduced them into the rice genome by Agrobacterium-mediated transformation. In this report, rice plants that contained single and simple insertions of T-DNA were analyzed in order to evaluate the gene-tagging efficiency. The 3'end of Ds was examined for putative splicing donor sites. As observed in maize, three splice donor sites were identified at the 3'end of the Ds in rice. Nearly 80% of Ds elements wered excised from the original T-DNA sites, when Ac cDNA was expressed under a CaMV 35S promoter. Repetitive ratoon culturing was performed to induce new transpositions of Ds in new plants derived from cuttings. About 30% of the plants carried at least one Ds that underwent secondary transposition in the later cultures. 8% of transposed Ds elements expressed GUS in various tissues of rice panicles. With cloned DNA adjacent to Ds, the genomic complexities of the insertion sites were examined by Southern hybridization. Half of the Ds insertion sites showed simple hybriodization patterns which could be easily utilized to locate the Ds. Our data demonstrate that the Ac/Ds mediated gene trap system could prove an excellent tool for the analysis of functions of genes in rice. We discuss genetic strategies that could be employed in a largee scale mutagenesis using a heterologous Ac/Ds family in rice.

We have previously isolated OsMADS4 gene that is a member of the class B MADS box genes from rice. In this study, another member of the class B MADS box genes was isolated from rice flower by the yeast two-hybrid screening method using OsMADS4 as bait. RNA blot analyses revealed that the clone, OsMADS16, was expressed in the second and third whorls, whereas the OsMADS4 transcripts were present in the second, third, and fourth whorls. These expression patterns of the OsMADS16 and OsMADS4 genes are very similar with those of AP3 and PI, the class B genes of Arabidopsis, respectively. In the yeast two-hybrid system, OsMADS4 interacted only with OsMADS16 among several rice MADS genes investigated, suggesting that OsMADS4 and OsMADS16 function as a heterodimer in specifying sepal and petal identities. We have also isolated OsMADS6 gene using OsMADS1 as a probe. Both are members of the AGL2 MADS family. Various MADS genes that encode for protein-protein interaction partners of the OsMADS6 protein were isolated by the yeast two-hybrid screening method. A majority of these genes belong to the AGL2 family. Sequence Homology, expression pattern, and ectopic expression phenotypes indicated that one of the interaction partners, OsMADS14, appears to be homologous to API, the class A MADS gene of Arabidopsis.

cDNAs for long- and short-chain acyl-CoA oxidases in fatty acid $\beta$-oxidation were isolated and were characterized their enzymatical and molecular properties. Both oxidases were exclusively localized in glyoxysomes, indicating that glyoxysomes can completely metabolize fatty acids to acyl-CoA by their cooperative action. In order to clarify the regulatory mechanisms underlying degradation of storage oil, we tried to obtain glyoxysome-deficient mutants of Arabidopsis. We screened 2,4-dichlorophenoxybutyric acid (2,4-DB) mutants of Arabidopsis which have defects in glyoxysomal fatty acid $\beta$-oxidation. Four mutants can be classified as carrying alleles at three independent loci, which we designated pedl, ped2, and ped3, respectively (where ped stands for peroxisome defective). The characteristics of these ped mutants are described.

Magnaporthe grisea (Hebert) Barr (anamorph: Pyricularia grisea) is a typical heterothallic Ascomycete and the causal agent of rice blast, one of the most destructive diseases on rice (Oryza sativa L.) worldwide. The interactions between cells of the pathogen and those of the host involve a complex of biological influences which can lead to blast disease. The early stages of infection process in particular may be viewed as a sequence of discrete and critical events. These include conidial attachment, gemination, and the formation of an appressorium, a dome-shaped and melanized infection structure. Disruption of this process at any point will result in failure of the pathogen to colonize host tissues. This may offer a new avenue for developing innovative crop protection strategies. To recognize and capture such opportunities, understanding the very bases of the pathogenesis at the cellular and molecular level is prerequisite. Much has been learned about environmental cues and endogenous signaling systems for the early infection-related morphogenesis in M. grisea during last several years. The study of signal transduction system in phytopathogenic filamentous fungi offers distinct advantages over traditional mammalian systems. Mammalian systems often contain multiple copies of important genes active in the same tissue under the same physiological processes. Functional redundancy, alternate gene splicing, and specilized isoforms make defining the role of any single gene difficult. Fungi and animals are closely related kingdoms [3], so inferences between these organisms are often justified. For many genes, fungi frequently possess only a single copy, thus phenotype can be attributed directly to the mutation or deletion of any particular gene of interest.

Plant cell culture is emerging to express bioactive foreign proteins because it has several advantages in that it is safe, economical, genetically stable and eukaryotic expression system comparing with other expression systems. However several limitations such as slow growth rate, low expression level and lack of well established down stream process need to be answered. As a preliminary approach to produce the immunologically interested molecules through the plant cell culture, we tested if granulocyte-macrophage colony stimulating factors (GM-CSFs) from both murine (mGM-CSF) and human (hGM-CSF) are produced as a biologically active form through plant cell culture. The murine and human GM-CSF genes were cloned into the plant expression vector, pBI121, and Ti-plasmid mediated transformation of tobacco leaves was conducted using Agrobacterium tumefaciens harboring both recombinant GM-CSF (rGM-CSF) genes. Cell suspension culture was established from the leaf-derived calli of transgenic tobacco plant. Northern blot analysis indicated the expression of the introduced mGM-CSF gene in both transgenic plant and cell suspension cultures. In addition, the biological activities of both murine and human GM-CSF from plant cell culture were confirmed by measuring the proliferation of the GM-CSF dependent FDC-PI and TF-1 cells, respectively.

Plastids, which are organelles unique to plant cells, bear their own genome that is organized into DNA-protein complexes (nucleoids). Regulation of gene expression in the plastid has been extensively investigated because this organelle plays an important role in photosynthesis. Few attempts, however, have been made to characterize the regulation of plastid gene expression at the chromosomal structure, using plastid nucleoids. In this report, we summarize the recent progress in the characterization of DNA-binding proteins in plastids, with special emphasis on CND41, a DNA binding protein, which we recently identified in the choloroplast nucleoids from photomixotrophically cultured tobacco cells. CND41 is a protein of 502 amino acids which consisted of a transit peptide of 120 amino acids and a mature protein of 382 amino acids. The N-terminal of the 'mature' protein has lysine-rich region which is essential for DNA-binding. CNA41 also showed significant identities to some aspartyl proteases. Protease activity of purified CND41 has been recently confirmed and characterized. On the other hand, characterization of accumulation of CND41 both in wild type and transgenic tobacco with reduced amount of CND41 suggests that CND41 is a negative regulator in chloroplast gene expression. Further investigation indicated that gene expression of CND41 is cell-specifically and developmentally regulated as well as sugar-induced expression. The reduction of CND41 expression in transgenic tobacco also brought the stunted plant growth due to the reduced cell length in stem. GA3 treatment on apical meristem reversed the dwarf phenotype in the transformants. Effects of CND41 expression on GA biosynthesis will be discussed.

Light-regulated translation of chloroplast mRNAs requires nuclear-encoded trans-acting factors that interact with the 5' untranslated region (UTR) of these mRNAs. A set of four proteins (60, 55, 47, and 38 kDa) that bind to the 5'-UTR of the psbA mRNA had been identified in C. reinhardtii. 47 kDa protein (RB47) was found to encode a chloroplast poly (A)-binding protein (cPABP) that specifically binds to the 5'-UTR of the psbA mRNA, and essential for translation of this mRNA, cDNA encoding 60 kDa protein (RB60) was isolated, and the amino acid sequence of the encoded protein was highly homologous to plants and mammalian protein disulfide isomerases (PDI), normally found in the endoplasmic reticulum (ER). Immunoblot analysis of C. reinhardtii proteins showed that anti-PDI recognized a distinct protein of 56 kDa in whole cell extract, whereas anti-rRB60 detected a 60 kDa protein. The ER-PDI was not retained on heparin-agarose resin whereas RB60 was retained. In vitro translation products of the RB60 cDNA can be transported into C. reinhardtii chloroplast in vitro. Immunoblot analysis of isolated pea chloroplasts indicated that higher plant also possess a RB60 homolog. In vitro RNA-binding studies showed that RB60 modulates the binding of cPABP to the 5'-UTR of the psbA mRNA by reversibly changing the redox status of cPABP using redox potential or ADP-dependent phosphorylation. Site-directed mutagenesis of -CGHC- catalytic site in thioredoxin-like domain of RB60 is an unique PDI located in the chloroplast of C. reinhardtii, and suggest that the chloroplast PDI may have evolved to utilize the redox-regulated thioredoxin like domain as a mechanism for regulating the light-activated translation of the psbA mRNA.

We have isolated more than a dozen cDNA clones corresponding to genes that were expressed in Arabidopsis leaves when they were kept in the dark. The nucleotide sequence analysis showed that some of the clones encoded proteins with significant homology to $\beta$-glucosidase (din2), branched-chain $\alpha$-keto acid dehydrogenase subunit E1$\beta$(din3), and another subunit E2 (din4), yeast RAD23 (din5), asparagine synthetase (din6), pre-mRNA splicing factor SRp35 (din7), phosphomannose isomerase (din9), seed imbibition protein (din10), and 2-oxoacid-dependent oxidase (din11). Accumulation of transcripts from din3,4,6 and 10 occurred rapidly after the plants were transferred to darkness. Transcripts from din2,9, and 11 could be detected only after 24 h of dark treatment. Inhibition of photo-synthesis by DCMU strongly induced the accumulation of transcripts from those genes, and application of sucrose to detached leaves suppressed the accumulation both in the dark and by DCMU. These observations indicate that expression of the genes is caused by sugar starvation resulted from the cessation of photosynthesis. We further showed that din2-encoded protein also accumulated in senescing leaves. Given these results, possible roles of din genes in leaves in the dark and senescing leaves are discussed.

In plants as well as in other multicellular organisms, programmed cell death plays essential roles in the abortion or formation of specific cells and tissues during development to organize the plant [11, 15, 18]. A typical example of developmentally programmed cell death in plants is the death during differentiation of tracheary elements which are components of vessels and tracheids, a water-conducting system. The programming of cell death during tracheary element differentiation has been revealed to be unique to plant cells by using the in vitro Zinnia mesophyll cell culture system. In particular, new biosynthesis of autolysis-related enzymes such as cysteine proteases and nucleases, their accumulation of the vacuole and the programmed collapse of the vacuole are essential to the death of tracheary elements and differ greatly from the process of the apoptotic cell death in animals.

To obtain direct evidence for thed involvement of C-AB13, a carrot (Daucus carota L.) homolog of VPI/Ab13, seed-specific transcription factor, in the acquisition of desiccation tolerance carrot non-embryogenic cells (NC) in which the C-AB13 gene was expressed ectopically was prepared. Non-transgenic NC, in which expression of C-AB13 was not detected, did not exhibit desiccation tolerance even after treatment with abscisic acid (ABA). In transgenic NC that expressed C-AB13, embryo-specific ABA-inducible genes (ECP genes) were expressed upon ABA-treatment. Furthermore, the transgenic NC became desiccation-tolerant upon ABA-treatment, but not tolerate desiccation without ABA-treatment. These results provide direct evidence for the involvement of C-AB13 in the ABA-induced acquisition of desiccation tolerance.

Oxidative stress is one of the major environmental stresses to plants. Reactive oxygen species (ROS) generated during metabolic processes damage cellular functions and consequently lead to cell death. Fortunately plants have in vivo defense system by which the ROS is scavenged by enzymes such as superoxide dismutase (SOD) and ascorbate peroxidase (APX). In attempts to understand the protection mechanism of plant against oxidative stress, we developed transgenic tobacco (Nicotiana tabacum cv. Xanthi) plansts thet expressed both SOD and APX in chloroplast using Agrobacterum-mediated transformation and evaluated their protection capabilities against methyl viologen (MV, paraquat) -mediated oxidative damage. Three double transformants (CAI, CA2, and CA3) expressed the chimeric CuZnSOD and chimeric APX in chloroplast, and one transformant (AM) expressed the chimeric APX and chimeric MnSOD in chloroplast. In addition, we obtained three lines of transformants (C/Al, C/A2, and A/C) that expressed the APX and SOD than control plants, and more resistant to oxidative stress caused by MV. TRansformants (C/A and A/C) overexpressing MnSOD, CuZnSOD and APX at the same time showed the highest resistance to MV-mediated oxidative stress among the transformants.

Somatic embryogendesis is one of good examples of the basic research for plant embryo development as well as an important technique for plant biotechnology. This paper describes the direct somatic embryogenesis from zygotic embryos of Panax ginseng is reversely related to normal axis growth of zygotic embryos by the experiment of various chemical treatments. Under the normal growth condition, the apical tips of embryo axis produced an agar-diffusible substance, which suppressed somatic embryo development from cotyledons. Although the cells of zygotic embryos were released from the restraint of embryo axis, various factors were still involved for somatic embryo development. Electron microscopic observation revealed that the ultrastructure of cells of cotyledon epidermis markedly changed before initiation of embryonic cell division, probably indicating reprogramming events into the cells embryogenically determined state. Polar accumulation of endogenous auxin or cell-cell isolation by plasmolysis pre-treatment is the strong inducer for the somatic embryo development. The cells for the process of somatic embryogenesis might be determined by the physiological conditions fo explants and medium compositions. Direct somatic embryos from cotyledons fo ginseng were originated eithrer from single or multiple cells. The different cellular origin of somatic embryos was originated either from single or multiple cell. The different cellular origin of somatic embryos was depended on various developmental stages of cotyledons. Immature meristematic cotyledons produced multiple cell-derived somatic embryos, which developed into multiple embryos. While fully mature cotyledons produced single cell-derived single embryos with independent state. Plasmolysis pretreatment of cotyledons strongly enhanced single cell-derived somatic embryogenesis. Single embryos were converted into normal plantlets with shoot and roots, while multiple embryos were converted into only multiple shoots. GA3 or a chilling treatment was prerequisite for germination and plant conversion. Low concentration of ammonium ion in medium was necessary for balanced growth of root and shoot of plantlets. Therefore, using above procedures, successful plant regeneration of ginseng was accomplished through direct single embryogenesis, which makes it possible to produce genetically transformed ginseng efficently.