• Food Science of Animal Resources
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• v.28 no.1
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• pp.99-104
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• 2008

A lactic acid bacterium showing growth inhibitory activity against Enterobacter sakazakii was isolated from bovine intestinal tracts. From biochemical and molecular biological studies, the isolate was identified and named as Enterococcus faecium JH95. This strain was resistant to kanamycin and streptomycin at a concentration of $100{\mu}g/mL$. E. faecium JH95 had high antimicrobial activity against food-borne pathogens such as Escherichia coli O157:H7, Listeria monocytogenes, Salmonella typhimurium, Staphylococcus aureus, and Clostridium perfrigens. The culture supernatant of this strain did not have antimicrobial activity. The culture broth of this strain failed to show the antimicrobial activity by heat treatment at $100^{\circ}C$ for 5 min or by pretense treatments for 2 hr. This result suggested that the putative antimicrobial substance produced by E. faecium JH95 is likely a protein which is not secreted into culture medium.

• Microbiology and Biotechnology Letters
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• v.23 no.2
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• pp.164-169
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• 1995

An endogenous cryptic plasmid, pBL1, which has been used to construct plasmid vectors for coryneform bacteria producing amino acids, was eliminated from Brevibacterium lactofermentum. The pBL1 was partially digested with Sau3AI and the resulting DNA fragments were subcloned into a suicide vector pEM1 which contains a kanamycin-resistant (km$^{r}$) gene. KM$^{r}$ B. lactofermentum transconjugants were obtained by conjugal transfer of the pEM1 derivatives containing pBL1 DNA fragments from Escherichia coli into B. lactofermentum. A km$^{r}$ transconjugant was analyzed to contain a plasmid pEB14, which occurred in vivo by homologous recombination between pBL1 and the conjugal-transferred plasmid. The pEB14 including the pEM1-derived km$^{r}$ gene was found to be lost concomitantly with km$^{r}$ phenotype, resulting in the construction of a pBL1-free strain of B lactofermentum. Based on transformation efficiencies and plasmid stability, the resultant pBL1- free strain is more useful than wild strain as a host cell for genetic manipulation. It could be concluded that foreign plasmid DNAs are efficiently isolated and analyzed from the pBL1-free strain because of the absence of endogenous pBL1 plasmid.

• Journal of Life Science
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• v.12 no.1
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• pp.67-76
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• 2002

The effect of transgene inactivation in T2, T3 and F2 generations was analyzed in progeny seedlings which had been generated by Agrobacterium (LBA4404/pBI121)-mediated transformation in Arabidopsis thaliana. In a system which investigated in the expression of $\beta$-glucuronidase(GUS)gene in kanamycin-resistant (ke $n^{R}$)seedlings, GUS inactivated seedlings were observed in 5 of 12 tested lines of T2 generation and the frequency of GUS inactivation was approximately 2.3%. Lines with multi-copies of T-DNA exhibited severe GUS gene inactivation with the frequency of 5.8% in T2 generation. In T3 generation lines exhibited GUS gene inactivation with the frequency of 1.3%. In contrast, inactivation increased dramatically up to 12.6% in multi-copy T-DNA line. A similar phenomenon was also found in F2 progeny from a transgenic line which had been crossed with wild-type Arabidopsis plant, WS-O (GUS gene inactivation frequency 9.9%). These results indicate that the foreign gene introduced into the plant was inactivated progressively in its transmission during subsequent generations and the transgenic line with multi-copies of T-DNA tended to show more increased inactivation.

• Korean Journal of Plant Tissue Culture
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• v.24 no.1
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• pp.63-69
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• 1997

Erythropoietin (EPO) is a glycoprotein that mediates the growth and differentiation of erythroid progenitors. In order to produce recombinant human erythropoietin in tobacco plant, the EPO genomic DNA (5.4 kb) was cloned into plant expression vectors, pBI$\Delta$GUS121, pBD$\Delta$GUS121 and pPEV-1, and introduced in Nicotiana tabacum (var. Xanthi) via Agrobacterium tumefaciens-mediated transformation. After selection on MS media containing kanamycin (Km), 10 Km-resistant plants were obtained per each construct. The correct integration of EPO genomic DNA in the genome of transgenic plant was confirmed by polymerase chain reaction (PCR). Northern blot showed that transcripts of 1.8 kb length were produced in leaves of the plants, but there was no difference of mRNA amount according to promoter number and 5'-untranslated sequence (UTS). The proteins obtained from leaves of transgenic plants were immunologically detected by Western blot using rabbit anti-human EPO polyclonal antibody. The expressed protein appeared as smaller band of apparent mass of 30 kDa as compared to the EPO protein from human urine (37 kDa), suggesting that the modification (glycosylation) system in tobacco plant might be different from that of mammalian cells.

• Korean Journal of Microbiology
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• v.46 no.3
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• pp.303-307
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• 2010

Acinetobacter baumannii 1625, a clinical isolate identified by Vitek and 16S rDNA sequence, showed an extended resistance to most ${\beta}$-lactams including imipenem, kanamycin, gentamicin, tobramycin, and cephalosporins of the third and fourth generations, and produced metallo-${\beta}$-lactamase (MBL) of IMP-1 type which is rare in Korea. The isolate contained a class 1 integron of about 2.5 kb in size and the integron included accA4 (aminoglycoside resistance gene), $bla_{IMP-1}$ (carbapenem resistance gene), and $bla_{OXA-2}$ (extended-spectrum ${\beta}$-lactam resistance gene) gene cassettes in order. The coexistence of IMP-1 type and OXA-2 type ${\beta}$-lactamase gene cassettes in an integron has not been reported in Korea. The transformed integron rendered the E. coli transformant resistant more than eight folds against imipenem, ampicilin, piperacillin, cefazolin, cefoperazone, and aztreonam comparing to the reference strain. This study clearly showed that the extended multi-drug resistance of A. baumannii 1625 was mainly due to the integron.

• Journal of Life Science
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• v.27 no.9
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• pp.1040-1046
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• 2017

We describe the isolation and characterization of six different intestinal microorganisms from the digestive tract of the cricket Gryllus bimaculatus. Based on 16S rRNA gene sequences, we obtained six isolates belonging to four different genera: Staphylococcus, Bacillus, Citrobacter, and Proteus. All the isolates were resistant to ampicillin. Ampicillin is an irreversible inhibitor of the enzymeetranspeptidase, which is needed to make bacterial cell walls. None of the isolates were resistant to kanamycin, which binds to the 30S subunit of the bacterial ribosome and then inhibits total protein synthesis. Gram staining was conducted, in addition to morphological classification under a microscope. Four grampositive isolates and two gram-negative isolates were detected. The gram-positive isolates were GL1 (round shaped, 2 am in diameter), GL2 (rod shaped, $2.5{\mu}m$ in length), GL3 (rod shaped, $2{\mu}m$ in length), and GL4 (round shaped, $1.5{\mu}m$ in diameter). The gram-negative isolates were GL5 (rod shaped, $2{\mu}m$ in length) and GL6 (rod-shaped, $2.5{\mu}m$ in length). Notably, two of the isolates, GL2 and GL4, secreted specific extracellular proteins. These were determined by MALDI-TOF-MS spectral analysis to be a 87 kDa collagenase, 56 kDa hypothetical protein, and 200 kDa hypothetical protein. The six isolates in this study could be used for various biotechnological applications and pest management, both in the field and in greenhouse systems. In addition, it would be interesting to determine the relationship between these isolates and their host.

• Microbiology and Biotechnology Letters
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• v.17 no.5
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• pp.447-453
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• 1989

In order to study the transfer of antibiotics resistance genes of the genetically cloned bacteria in water environments, DK1 strain, which is resistant to kanamycin (Km), chloramphenicol (Cm), streptomycin (Sm), and sulfadiazine (Su), was selected from the Gram-negative bacterial isolates from wastewater. One of 4 plasmids harboured in the DK1 strain was found to possess Km$^{${\gamma}$}$Cm$^{${\gamma}$}$ gene and be about 68 kb in size, and it was designated as pDK101. The plasmid of pDK101 was also found to have 16, 32, and 6 restriction sites for EcoRI. .PstI, and SalI, respectively. From the digestion fragments of pDK101 plasmid and pKT230 used as a vector by EcoRI restriction endonuclease, pDT309 and pDT529 were constructed as chimeric plasmids which possess Km$^{${\gamma}$}$Cm$^{${\gamma}$}$ gene and are 30.9 and 52.9 kb in size, respectively. When the chimeric plasmids were trasformed into E. coli C600 or E. coli HB101, transformants of DKC601, DKC602, DKH102, and DKH103 were obtained as cloned bacterial cells. The Km$^{${\gamma}$}$Cm$^{${\gamma}$}$ genes were well expressed in those cloned cells and the chimeric plasmids were clearly detected in the cloned cells of DKC601 and DKH103.

• Korean Journal of Food Science and Technology
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• v.38 no.2
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• pp.295-303
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• 2006

To provide microbial information for the safety of agricultural production, the presence of enterotoxin genes and antibiotic susceptibility of 14 isolated Staphylococcus aureus (11.7%) strains were investigated using PCR-based methods and disk diffusion method, respectively. Among enterotoxin-encoding genes, sea was detected from two isolates (14.3%), sea and sed genes were co-detected from three isolates (21.4%), and sea, sed, and see genes in seven isolates (50.0%), whereas seb, sec, and tsst were not detected in any isolate. Nine (64.3%), eight (57.1%), six (42.9%), two (14.3%), and one (7.2%) isolates were resistant to penicillin, novobiocin, amphicillin, erythromycin and oxacillin, and doxycycline and kanamycin, respectively. Methicilline-resistant S. aureus was found in roller of B farm and in hydroponic solution of D farm.

• Journal of Food Hygiene and Safety
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• v.16 no.3
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• pp.159-167
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• 2001

Staphylocorccus aureus is gram positive, facultatively anaerobic, non-sporulative coccus, and positive for coagulase and DNase. The food-poisoning outbreak of Staphylococcus aureus increases in the world, and third occurrence happened in our country. Of 105 isolates (25.4%) obtained 413 focal samples of food-poisoning suspicious patients. In those cases, the enterotoxins were detected from a total of 45 isolates (42.9%), 9 isolates(20.0%) were A type, 33 isolates (73.3%) were H types, 2 isolates (4.4%) were G type and 1 isolate was a I type enterotoxin. Among the isolates possessing staphylococcal enterotoxins, 29 isolates had H type only(64.4%), 5 isolates had A type only and 4 isolates had both A and H type. Two isolates had G type only and 1 isolate had I type only. In the antibiotic susceptibility, 48 isolates (46%) had at least one antibiotic resistance among 105 isolates, 34 isolates (70.8%) were resistant to penicillin. 1 isolate (2.1%) to ampicillin, 3 isolates (6.3%) to erythromycin and kanamycin. Seven were resistant to more than two antibiotics and especially 1 isolate was resistant to penicillin-ampicillin-nitrofurantoin.

• Korean Journal of Microbiology
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• v.26 no.4
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• pp.305-311
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• 1988

The spectinomycin resistant strain of slow growing R. japonicum R-168 was selected to be participated in conjugation with E. coli WA803/pGS9. Tn5 was introduced from suicide vector pGS9 into R. japonicum R-168 $spr^{r}$ chromosome at the frequency of $1.0\times 10^{-5}-5.0\times 10^{-7}$ and the transconjugante were selected on the yeast extract-mannitol plate containing kanamycin ($50{\mu}$g/ml) and spectinomycin ($100{\mu}$g/ml) after 8-9 days incubation. All transconjugants we tested were found to contain Tn 5 DNA on their genome, which was confirmed by Southern hybridization experiments. R. japonicum RNa75, which had been selected through plant test, was found to be defective in symbiotic nitrogen fixing ability and the production of leghemoglobin in soybean nodules formed by the inoculation of this mutant. In addition, this mutant strain hardly developed nitrogenase activity asymbiotically in contrast with the wild type strain, indicating that some nitrogen fixing gene might be blocked in this strain and the production of leghemoglobin could be decreased by the interference in nitrogen fixing genes.