• Journal of Plant Biology
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• v.38 no.1
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• pp.19-24
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• 1995

The vir genes expression of Ti plasmid is induced by a family of related phenolic compounds. We investigated the effects of various phenolic compounds, Ti plasmids and hosts on the expression of the vir genes in the same type of octopine Ti plasmids, pTiKU12, pTiAch5 and pTiA6. The vir gene induction of pTiKU12 was remarkably stimulated by p-coumaric acid in relation to acetosyringone, but those of pTiAch5 and pTiA6 were more stimulated by acetosyringone than by p-coumaric acid. The effect of phenolic compound on the vir gene induction was different according to the kind of Ti plasmids. Also, the vir gene expression of A. tumefaciens KU913, which has pTiKU12 was about 6.2 times as much as that of A. tumefaciens KU915, which has pTiKU12 in KU12 host, in the presence of ferulic acid. But no difference was shown in the presence of p-coumaric acid. The vir gene induction abilities of phenolic compounds are different according to the kinds of phenolic compounds, Ti plasmids and hosts.

• Research in Plant Disease
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• v.15 no.2
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• pp.77-82
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• 2009

The roots of grapevine in the field in which the crown gall was occurred severely in Chungbuk province were collected and Agrobacterium spp. were isolated from the roots using the selective media. The selected 13 isolates were identified as A. tumefaciens with fatty acid analysis using MIDI system, nucleotide sequence of 16S rDNA, biochemical characteristics, and PCR with the species-specific primers. A. vitis, a pathogen of crown gall disease of grapevine was not isolated from the roots. All of the isolates did not show pathogenicity on the tomato seedlings and the stem and root of grapevine. Eric-PCR showed that DNA band patterns of the root isolates were a little more similar to A. tumefaciens than A. vitis. However, overall similarity between the root isolates and the pathogenic strains of A. tumefaciens and A. vitis was low by rep-PCR. These results suggest that a pathogen causing crown gall in grapevine in Chungbuk province may transmitted through the seedlings rather than via soil or roots.

• Korean Journal of Microbiology
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• v.27 no.2
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• pp.92-97
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• 1989

the vector, pGUR19, for Rhizobium gene manipulation, was constructed by combining the replication and mobilization function of indigenous multicopy plasmid from Acacia(Robinia pseudoacacia L.) Rhizobia sp86 with E. coli cloning vehicle, pBR322. The vector could be efficiently mobilized by RP4 tra function incorporated into chromosome of E. coli named SM10 and efficiently transferred to various gram negative hosts including Rhizobium and Afrobacterium by transformation. Mobilization frequency of the constructed vector was ranged from $1.2\times 10^{-2}$ (E.coli HB 101) to $4.6\times 10^{-4}$ (A. tumefaciens 15955) and transformation frequency was ranged from $5.4\times 10^{-7}$(E. coli HB101) to $1.2\times 10^{-10}$ (A. tumefaciens 15955). The vector, pGUR19, was stably replicated and maintained in a variety of Rhizobium and Agrobacterium.

• Journal of Microbiology and Biotechnology
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• v.34 no.5
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• pp.1178-1187
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• 2024

Cordyceps militaris is a significant edible fungus that produces a variety of bioactive compounds. We have previously established a uridine/uracil auxotrophic mutant and a corresponding Agrobacterium tumefaciens-mediated transformation (ATMT) system for genetic characterization in C. militaris using pyrG as a screening marker. In this study, we constructed an ATMT system based on a dual pyrG and hisB auxotrophic mutant of C. militaris. Using the uridine/uracil auxotrophic mutant as the background and pyrG as a selection marker, the hisB gene encoding imidazole glycerophosphate dehydratase, required for histidine biosynthesis, was knocked out by homologous recombination to construct a histidine auxotrophic C. militaris mutant. Then, pyrG in the histidine auxotrophic mutant was deleted to construct a ΔpyrG ΔhisB dual auxotrophic mutant. Further, we established an ATMT transformation system based on the dual auxotrophic C. militaris by using GFP and DsRed as reporter genes. Finally, to demonstrate the application of this dual transformation system for studies of gene function, knock out and complementation of the photoreceptor gene CmWC-1 in the dual auxotrophic C. militaris were performed. The newly constructed ATMT system with histidine and uridine/uracil auxotrophic markers provides a promising tool for genetic modifications in the medicinal fungus C. militaris.

• Journal of Microbiology and Biotechnology
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• v.14 no.4
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• pp.822-828
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• 2004

Nopaline-type Agrobacterium tumefaciens strain C58 cannot utilize octopine (Oct) as the sole carbon and nitrogen sources. This strain harbors two plasmids; a virulent plasmid, pTiC58, and a megaplasmid, pAtC58. From strain NT1, which is a derivative of C58 harboring only pAtC58, we isolated spontaneous mutants that utilize Oct as the sole nitrogen source. These Oct-catabolizing mutants, however, could not utilize the opine as the sole carbon source. In contrast, strain UIA5, a plasmid-free derivative of C58, could not give rise to such mutants. The mutations isolated from NT1 were mapped to socR in pAtC58, which is a negative regulator of the soc operon responsible for the uptake and catabolism of an Amadori opine, deoxyfructosyl glutamine (Dfg). A derivative of UIA5 carrying a clone of the soc operon with a transposon inserted in socR also utilizes Oct as the sole nitrogen source. However, UIA5 harboring the operon with mutations in each of the structural genes in the soc operon, socA, B, C, and D, lost the ability to generate spontaneous Oct-utilizing mutants, suggesting that soc genes in pAtC58 are required for the utilization of Oct as a nitrogen source, and that derepressed expression of these genes allows cells to utilize Oct. In contrast, Oct-catabolizing mutants derived from C58, which grew using Oct as the sole nitrogen source, could also utilize the opine as the sole carbon source. These mutants did not carry any detectable mutations in socR or the region upstream to the gene in pAtC58, suggesting that mutations occurring elsewhere in the genome, most likely in pTiC58, allow the uptake and catabolism of the opine.

• Journal of Microbiology and Biotechnology
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• v.26 no.11
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• pp.1924-1932
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• 2016

Mycophenolic acid (MPA) is an antibiotic produced by Penicillium brevicompactum. MPA has antifungal, antineoplastic, and immunosuppressive functions, among others. ${\beta}-Hydroxy-{\beta}-methylglutaryl-CoA$ (HMG-CoA) lyase is a key enzyme in the bypass metabolic pathway. The inhibitory activity of HMG-CoA lyase increases the MPA biosynthetic flux by reducing the generation of by-products. In this study, we cloned the P. brevicompactum HMG-CoA lyase gene using the thermal asymmetric interlaced polymerase chain reaction and gene walking technology. Agrobacterium tumefaciens-mediated transformation (ATMT) was used to insert a mutated HMG-CoA lyase gene into P. brevicompactum. Successful insertion of the HMG-CoA lyase gene was confirmed by hygromycin screening, PCR, Southern blot analysis, and enzyme content assay. The maximum MPA production by transformants was 2.94 g/l. This was 71% higher than wild-type ATCC 16024. Our results demonstrate that ATMT may be an alternative practical genetic tool for directional transformation of P. brevicompactum.

• Journal of Microbiology and Biotechnology
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• v.29 no.2
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• pp.230-234
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• 2019

Currently, the genetic modification of Aspergillus oryzae is mainly dependent on protoplast-mediated transformation (PMT). In this study, we established a dual selection marker system in an industrial A. oryzae 3.042 strain by using Agrobacterium tumefaciens-mediated transformation (ATMT). We first constructed a uridine/uracil auxotrophic A. oryzae 3.042 strain and a pyrithiamine (PT)-resistance binary vector. Then, we established the ATMT system by using uridine/uracil auxotrophy and PT-resistance genes as selection markers. Finally, a dual selection marker ATMT system was developed. This study demonstrates a useful dual selection marker transformation system for genetic manipulations of A. oryzae 3.042.

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

• Journal of Life Science
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• v.11 no.4
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• pp.316-320
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• 2001

Sonication tremendously improves the efficiency of Agrobacterium infection by introducing small and uniform fissures and channels throughout the targeted tissue. Using shoot tips of cotton as explants, the effect of sonication treatment and virulence genes in Agrobacterium tumefaciens on transformation efficiency was investigated. The pat gene which encodes resistance to the herbicide, glufosinate, was used as a selectable marker. Transformation efficiency was evaluated on th basis of survival rates of cocultivated shoot tips on selection medium containing 2.5 mg/l gulfosinate-ammonium(ppt) adn 25. mg/l Clavamax. Sonication from 5 to 15 second has a positive effect on shoop tip survival. However, whil virE as well as virG or vir GN54D showed an enhancement in transformation efficiency, virE,. virG resulted in the most significant enhancement. Overall, the combination of additional virG/virE gene and sonication treatment resulted in the most significant increase in transformation efficiency.

• Korean Journal of Microbiology
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• v.10 no.1
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• pp.41-50
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• 1972

The tomato plant, Lycopersicon esculentum Mill, was inoculated with tumor inducing strain, $A_6K_1$, of Agrobacterium tumefaciens and its produced tumors were examined with the electron microscope. A number of bacteria are usually detected in the intercellular region of the host plant, and it is observed that the host cytoplasm is readily destroyed in the region where the bacterial invasion occurred. Some of the bacteria in the host tissues are enclosed with the single unit membranes, in other locations lots of bacteroids were examined and the bacterial lysis is generally observed in those bacteroids. The bacterial movement in the tumor tissue and some peculiar relationships between the pathogens and the host plant are discussed.