• Journal of Plant Biotechnology
• /
• v.34 no.4
• /
• pp.285-291
• /
• 2007

This study describes conditions for the mass production of activation-tagged mutant hairy root lines of ginseng by cocultivation with Agrobacterium rhizogenes. Because it is not currently possible to produce progeny from transgenic ginseng, a loss-of-function approach for functional genomics cannot be appliable to this species. A gain-of-function approach is alternatively the choice and hairy root production by cocultivation of A. rhizogenes would be most practical to obtain a large number of mutants. Various sources of explants were subjected to genetic transformation with various strains of A. rhizogenes harboring the activation-tagging vector pKH01 to determine optimum conditions for the highest frequency of hairy root formation on explants. Petiole explants cocultivated with A. rhizogenes R1000 produced hairy roots at a frequency of 85.9% after 4 weeks of culture. Conditions for maximum growth or branching rate of hairy roots were also investigated by using various culture media. Petiole explants cultured on half strength Schenk and Hildebrandt medium produced vigorously growing branched roots at a rate of 2.6 after 4 weeks of culture. A total of 1,989 lines of hairy root mutants were established in this study. These hairy root lines will be useful to determine functions of genes for biosynthesis of ginsenosides.

• Research in Plant Disease
• /
• v.18 no.4
• /
• pp.268-276
• /
• 2012

Broad bean wilt virus 2 (BBWV2), genus Fabavirus, subfamily Comovirinae, family Secoviridae, causes damage in many economically important horticultural and ornamental crops. Sequence alignments showed several conserved sequences in 5' non-coding regions (5' NCRs) of RNA 1 and RNA 2 in all BBWV2 strains characterized so far. Based on this observation, we generated a hpRNA construct (pIR-BBWV2) harboring an inverted repeat containing a 210 bp cDNA fragment homologous to 5' NCR portion of BBWV2 RNA 1 to investigate the silencing potential for its ability to interfere with a rapidly replicating BBWV2. Agrobacterium-mediated transient expression of the IR-BBWV2 had a detrimental effect on BBWV2 infection, showing no distinct symptoms in non-inoculated leaves of the agroinfiltrated Nicotiana benthamiana plants. BBWV2 genomic RNAs were not detected by RT-PCR from tissues of both the inoculated leaves and upper leaves of the agroinfiltrated plants. Accumulation of virus-derived small interfering RNAs was detected in the inoculated leaf tissues of N. benthamiana plants elicited by transient expression of IR-BBWV2 indicating that RNA silencing is responsible for the resistance to BBWV2.

• KSBB Journal
• /
• v.5 no.3
• /
• pp.247-253
• /
• 1990

These studies were carried out to obtain the transformant from carrot cells by using binary vector pGA472 with NPT II gene to confer kanamycin resistance in the plant cells. The binary vector pGA472 was mobilized from E. coli MC1000 into A. tumefaciens strains isolated in the Korea, C23-1. K29-1, and disarmed Ti-plasmid PC2760, and A28l using a tri-parental mating method with E. coli HB101/pRK2013. Transconjugants, C23-1/pGA472, K29-1/pGA472, PC2 760/pGA472 and A28l/pGA472 were obtaind on the minimum AB media containing tetracycline and kanamycin, were comfirmed to hold the Ti-plasmid and pGA472 binary vector on the 0.7% agarose gel. Transformed carrot calli were initiated on the MS media supplemented with l00$\mu\textrm{g}$/ml kanamycin and 250$\mu\textrm{g}$/ml carbenicillin after co-cultivation of carrot explant and transconjugant Agrobacteria. Selected callus was grown vigouousley for subculture on the medium containing 100$\mu\textrm{g}$/ml kanamycin, thus indication that the selected callus was transformed with NPT II gene.

• The Korean Journal of Ecology
• /
• v.21 no.5_3
• /
• pp.703-712
• /
• 1998

Physicochemical factors, microbial population size and the properties of the bacterial isolates were estimated to find out the nature of soil ecosystem of Mt. Nam. Samples were obtained from the surface layer of soils on which specific plant community is developed. Average content of moisture and organic matter of the soils were 21.6% and 17.3%, respectively. These values were similar to those of developing forest soils, but were slightly lower than those of climax ecosystem such as Piagol in Mt. Chiri. Chiri. Content of phosphate was higher than those of other forest soils. The population size of soil bacteria ranged from 27.4 to 195.8 ${\times}\;10^5$ CFU/g. duy soil, and the size is somewhat dependent on the moisture and oranic matter content of soils. A large number of bacteria were able to decompose macromolecules such as starch, elastin and gelatin. Bacterial species composition of each soil was comparatively simple. Pseudomonas, Agrobacterium, Flavobacterium and Xanthomonas which are Gram-negative short rods were widely distributed in the forest soils. The endospore forming Bacillus species were also the main constituents of the soil microflroa. Actinomycetes were widely distributed in the forest soils, but the distribution pattern varied in each site. Most of the actinomycetes were also able to decompose organic macromolecules. The rate of resistant actinomycete strains to antibiotics and heavy metals were lower than those from cultivated soils, but higher than those from well-preserved forest soils. Antibiosis pattern of the actinomycete isolates was similiar to the resistance pattern. This means the forest soils of Mt. nam was somewhat interferred by artificial behabiour.

• Korean Journal of Soil Science and Fertilizer
• /
• v.35 no.2
• /
• pp.118-126
• /
• 2002

To evaluate the correlations of microbial populations with soil healthiness and crop production and establish the criteria for microbial population of soil types. We analyzed the microbial community structure of 13 soils which were different in physical and chemical properties and cultivation methods. According to the analysis of microbial population suing the dilution plate method, the large differences of the microbial population structures among soil types were shown: aerobic bacteria $2-27{\times}10^6$, fluorescent Pseudomonas $1-1,364{\times}10^5$, Gram negative bacteria $1-126{\times}10^4$, and mesophilic Bacillus $1-110{\times}10^5$. The density of Gram negative bacteria was highest on red pepper cultivating soils (sample no. 4 and 6) of Umsung and Gesan, Chungbuk, and the density of the fluorescent Pseudomonas was highest on greenhouse soil (sample no. 7) of Jinju, Kyungnam. The crop productivity of three soils was high as compared with those of other soils. It was supposed that the density of fluorescent Pseudomonas and mesophilic Bacillus were correlated with the incresed crop production. By MIDI analysis, 579 strains isolated from 13 soils composed of a variety of microbes including 102 isolates of Agrobacterium, 112 isolates of Bacillus, 32 isolates of Pseudomonas, 44 isolates of Kocuria, and 34 isolates of Pseudomonas. Among the 624 isolates of Gram negative bacteria, Pseudomonas including P. putida and p. fluorescens occupied the highest density (51%), and Stenotrophomonas maltophilia and Burkholderia cepacia also appeared at high density. From RAPD analysis, the fluorescent Pseudomonas strains isolated from 13 soil types showed a high level of strain diversities and were grouped into 2 - 14 patterns according to soil types. Many of unknown bacteria were recovered from the paddy soil, and needed to be further characterized on the molecular basis.

• Journal of Korean Society of Forest Science
• /
• v.87 no.3
• /
• pp.334-340
• /
• 1998

This research was conducted to identify plant regulatory regions by gene tagging method. A promoterless GUS coding sequence was introduced to Populus nigra ${\times}$ P. maximowiczii via Agrobacterium strains(LBA4404/EHA101), and putative transgenic poplars were selected by culturing on medium containing G418($60mg/{\ell}$) and by GUS assay. Among them one positive plant was to amplify the native sequences flanking to the introduced GUS gene in plant genome by inverse PCR method and from this 730 by DNA product was obtained. After subcloning and sequencing, it has 88% homology to the Eucalyptus gunnii CAD(cinnamyl alcohol dehydrogenase) gene. The GUS gene fused with the putative promoter reinserted into poplar leaves by particle bombardment method to test the funtional promoter activity. Upon staining with X-gluc, many blue spots appeared on the leaf segments bombarded by the chimeric gene 2-3 days, thus the isolated DNA fragment contain some possible coding region as well as a putative regulatory sequences of poplar CAD gene.

• Journal of Plant Biotechnology
• /
• v.33 no.4
• /
• pp.271-276
• /
• 2006

Hypocotyl explants of Chinese cabbage (us. 'Jeong Sang' and 'Seoul') produced adventitious shoots on Murashige and Skoog (MS) basal medium supplemented with 4mg/L $AgNO_3$, 5 mg/L acetosyringone, 4 mg/L 6-benzyladenine and 3mg/L alpha-naphthaleneacetic acid (SI) after cocoultivation with strains of Agrobacterium tumefaciens (LBA4404) harboring the pCAMBIA1301 and the $_PPTN290$ containing hygromycin-resistance gene and paromomycin-resistance gene as a selectable marker genes, respectively. There was a significant difference in the frequency of transgenic plants depending on antibiotics and cultivars used. Paromomycin was better than hygromycin, and cultivar 'Jeong-sang' was higher than 'c.v. Seoul' in the frequency of transgenic plants. In particular, the highest frequency (0.70%) of transgenic plants was obtained from selection medium (SI) containing 100mg/L paromomycin in c.v., 'Jeong-sang' GUS positive response were obtained 9 plants and 3 plants from the cultivars, 'Jeong-sang' and 'Seoul', respectively. They were grown to maturity in a greenhouse and normally produced $T_1$ seeds. GUS histochemical assay for progeny $(T_1)$ revealed that the transgenes were expressed in the plant genome.

• Research in Plant Disease
• /
• v.13 no.2
• /
• pp.98-103
• /
• 2007

Grapevines will experience various types of winter damage. Some winter damages are caused by mechanical injury, freezing temperatures or poor vine vigor. This research was conducted to find out the appropriate control methods through selection of resistant rootstocks and improvement of overwintering methods for the control of crown gall disease on 'Kyoho' grape. The crown gall symptoms were not found when three stock plants of grapevine SO4, 5BB and 3306 were inoculated with $10^4cfu/ml$ of Agrobacterium vitis strains (YK2823, YK3312, LMG259, HKA234). But when they were inoculated with higher concentration $(10^6 cfu/ml)$ of A. vitis, irrespective of stocks plants, crown galls were formed all of them and the gall size was much smaller than that of kyoho. Three stock plants were selected as resistant based on above mentioned. Covering trunks and branches with rice straw and insulating coverlet was the most effective method for prevention of crown gall disease. This treatment minimized the ambient temperature changes on grapevine trees during winter season to $9.6^{\circ}C$ and the normal plant growth was due to the absence of freezing injury.

• Proceedings of the Korean Society of Tobacco Science Conference
• /
• 1997.10a
• /
• pp.134-152
• /
• 1997

Plant viruses of tobacco including tobacco mosaic virus (TMV) and potato virus Y (PVY) cause severe economic losses in leaf-tobacco production. Cultural practices do not provide sufficient control against the viruses. Use of valuable resistant cultivars is most recommendable for the control of the viruses. However, conventional breeding programs are not always proper for the development of virus-resistant plants mostly owing to the frequent lack of genetic sources and introduction of their unwanted properties. Therefore, we tried to develop virus-resistant tobacco plants by transforming commercial tobacco cultivars, NC 82 and Burley 21, with coat protein (CP) or replicase (Nlb) genes of TMV and PVY necrosis strain (PVY-VN) with or without untranslated region (UTR) and with or without mutation. Each cDNA was cloned and inserted in plant expression vectors with 1 or 2 CaMV 35S promotors, and introduced into tobacco leaf tissues by Agrobacterium tumefaciens LBA 4404. Plants were regenerated in kanamycin-containing MS media. Regenerated plants were tested for resistance to TMV and PVY In these studies, we could obtain a TMV-resistant transgenic line transformed with TMV CP and 6 genetic lines with PVY-VN cDNAs out of 8 CP and replicase genes. In this presentation, resistance rates, verification of gene introduction in resistant plants, stability of resistance through generations, characteristics of viral multiplication and translocation in resistant plants, and resistance responses relative to inoculum potential and to various PVY strains will be shown. Yield and quality of leaf tobacco of a promising resistant tobacco line will be presented.