• Korean Journal of Microbiology
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• v.28 no.3
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• pp.219-228
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• 1990

A kanamycin resistance($Km^r$) gene was studied for its transfer in natural freshwater environments by using the natural bacterial isolate(M1) of DK1 and the DKC601 strain, $Km^r$ plasmid of which was genetically engineered from the NI strain. The transfer frequency ofthe $Km^r$ gene and rearrangement of the $Km^r$ plasmid were compared between the gnetically engineered microorganism(GEM) and the NI parental strain by conjugation with the same recipient strain. The transfer frequency of the $Km^r$ gene was about $9.1\times 10^{-12}-1.8\times 10^{-11}$ in both the GEM and NI strains at 5 to $10^{\circ}C$, but the frequency of the NI was about 10 times higher than that of the GEM at 20 to $30^{\circ}C$. The $Km^r$ plasmid in the transconjugants obtained by conjugation of the NI with the MY1 strain as a ricipient showed alot of rearrangement, but the $Km^r$ plasmid transferred from the GEM was stable without alteration of its size. When the MT2 strain was used as a recipient, however, such a rearrangement of the $Km^r$ plamid was observed in the transconjugants obtained from the GEM as well as the NI strain. In those transconjugants obtained from different mating pairs and water environments, the plasmid were appeared to decrease in their number as the period of conjugation time was prolonged, but only the $Km^r$ plasmid transferred from the GEM kept having its size of 52kb. Therefore, the $Km^r$ gene was transferred at the same rate from the GEM and NI strains in natural freshwater environment, but the gene of the GEM strain was more stable than the NIduring conjugation and the $Km^r$ plasmid was rearranged by changing the recipient strain for conjugation in any water environments.

• Journal of fish pathology
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• v.6 no.1
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• pp.57-64
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• 1993

The properties and DNA structures of R plasmids differ depending on the species of the fish-pathogens Aeromonas hydrophila, A. salmonicida, Edwardsiella tarda, Enterococcus seriolicida, Pasteurella piscicida and Vibrio anguillarum. However, some R plasmids with the same resistance markers in similar DNA structures were found in A. hydrophila and E. tarda, as well as in A. hydrophila and A. salmonicida. R plasmids from V. anguillarum were classified into three groups according to their DNA structures. The first group was detected before 1977, the second from 1980 to 1983, and the third from 1989 to 1991. R plasmids have been retained within P. piscieida having the same DNA structures and detected at various locations and times. E. seriolicida strains carrying the same R plasmids, which were encoded with resistance to macrolide antibiotics(MLs), lincomycin(LIM), and TC, and to MLs, LIM, and CP. were distributed in yellowtail farms in various districts. The chloramphenicol-resistance(cat) gene of the R plasmids of P. piscicida was classified as CAT type I. The cat of the R plasmids of E, tarda. A. salmonicida was classified as type II. The cat of R plasmids of V. anguillarum was classified into two types. One type detected before 1977, was classified as CAT IV and the other type, detected after 1980, was classified as CAT II. Tetracycline-resistance (tet) V. anguillarum, isolated before 1977 and after 1981, was classified as Tet B and Tet G, respectively. The class D tet gene was widely distributed in R plasmids from fish-pathogens A. hydrophila, E. tarda, P. piscicida, and also V. anguillarum isolated after 1989.

• Korean Journal of Microbiology
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• v.32 no.3
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• pp.186-191
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• 1994

The origin of the leading strand replication (ori) and of lagging strand replication (M-O) of R-plasmid pSBK203 was identified and its base sequence was determined. About 50 bp of ori sequence residues overlapped with the structural gene of rep. Sequence comparison reveals that pSBK-ori shares obvious identities with those of pT181 family and consists of two regions, one is conserved and the other is variable region. Of two palindrome sequence located one after another in upstream region of rep gene, palA' instead of palA which shares sequence homology with diverse family of plasmids such as pOX6, pC194, and pE194 seems to act as a signal for conversion of primarily replicated ssDNA to dsDNA (minus origin (M-O)).

• Applied Biological Chemistry
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• v.28 no.3
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• pp.149-155
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• 1985

590 strains of Rhizobia were isolated from root nodules of the legumes collected at 223 sites in Korea. According to their host specificities they were classified into R. japonicum(218 strains), R. phaseoli(101 strains), R. trifolii(97 strains), R. meliloti(4 strains), R. leguminosarium(1 strain), Rhizobium species(101 strains), and unidentified species(159 strains). 3 potent strains R-138, R-168, and R-214 of R. japonicum have been selected based on the infectivity to soybean cultivar and effeciency of nitrogen fixation. It was observed that the fast-growing strains of R. japonicum contained 1 to 4 plasmids of M.W. of 35-300 Md. However, plasmids were hardly detected for the slow-growing strains.

• Journal of fish pathology
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• v.16 no.1
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• pp.61-68
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• 2003

MICs of 8 chemotherapeutic agents against forty streptococcal isolates were determined. These strains were isolated from diseased olive flounder, Paralichthys olivaceus, and showed 2-5 multiple drug resistance against different antibacterial agents including ampicillin, doxycycline (DOXY), erythromycin, florfenicol, flumequine, norfloxacin, oxolinic acid, and oxytetracycline (OTC). In conjugation experiment, we found transferable R plasmids carrying OTC and DOXY resistance determinant in 3 drug resistance strains analyzed. Six out of 40 isolates showed positive signal in colony hybridization with the R plasmid DNA (pST9) as a probe. It suggests that other types R plasmid different from pST9 is also involving in multiple drug resistance of streptococci isolated from olive flounder.

• BMB Reports
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• v.37 no.3
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• pp.351-355
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• 2004

Separate protocols are commonly used to prepare plasmid DNA, chromosomal DNA, or total RNA from E. coli cells. Various methods for the rapid preparation of plasmid DNA have been developed previously, but the preparation of the chromosomal DNA and total RNA are usually laborious. We report here a simple, fast, reliable, and cost-effective method to extract total nucleic acids from E. coli by direct lysis of the cells with phenol. Five distinct and sharp bands, which correspond to chromosomal DNA, plasmid DNA, 23S rRNA, 16S rRNA, and a mixture of small RNA, were observed when analyzing the prepared total nucleic acids on a regular 1-2% agarose gel. The simple and high-quality preparation of the total nucleic acids in a singe tube allowed us to rapidly screen the recombinant plasmid, as well as to simultaneously monitor the change of the plasmid copy number and rRNA levels during the growth of E. coli in the liquid medium.

• Korean Journal of Microbiology
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• v.29 no.6
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• pp.408-415
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• 1991

TOL plsmid size of Flavobacterium odoratum SUB53 was estimated as 83 Md and the optimum concentration of m-toluate degradation by TOL plasmid was 5 mM. $H_{2}$ production by Rhodopseudomonas sphaeroides KCTC1425 was largely dependent on nitrogenase activity and showed the highest at 30 mM malate with 7 mM glutamate as nitrogen source. Nitrogenase activities were inhibited by 0.3 mM $NH_{4}^{+}$ions, to be appeared the decrease of $H_{2}$ production. Conjugation of TOL plasmids from F. odoratum SUB53 and Pseudomonas putida mt-2 to R. sphaeroides showed the optimum at the exponential stage of recipient cells in presence of helper plasmid pRK2013. According to the investigation of catechol-1,2-oxygenase (C-1, 2-O) and catechol-2,3-oxygenase (C-2,3-O) activities of R. sphaeroides C1 (TOL SUB53) and C2 (TOL mt-2), the gene for C-2,3-O is located on TOL plasmid and gene for C-1, 2-O on the chromosome of R. sphaeroides. m-Toluate was biodegraded by TOL plasmid in R. sphaeroides C1 and C2, presumably to be produced $H_{2}$ gas from the secondary metabolites of m-toluate.e.

• Korean Journal of Microbiology
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• v.25 no.1
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• pp.46-51
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• 1987

In order to increase the production of tryptophan by maximizing expression of recombinant trp plasmid, Klebsiella pneumoniae KC 105(pheA tyrA trpE trpR tyrR) was genetically modified. KC 107, inosine monophospate(IMP) auxotroph from KC 105 and KC 108, histidine(His) auxotroph from KC 107 were also derived respectively to increase phosphoribosylpyrophosphate(PRPP) production which is required for tryptophan biosynthesis. From KC 107 phosphoribosylpyrophosphate consumption which is required for tryptophan biosynthesis. From KC 107 and KC 108, KC 109 and KC 110, both arginine auxotrophs were derived respectively. To investigate the expression of recombinant trp plasmid in the selected K. pneumoniae mutants, the auxotrophic mutants were transformed with recombinant trp plasmids pSC 101-$trpE^{FBR}$, pSC 101-trpL(.DELTA.att) $trpE^{FBR}$ (pSC 101-trp-AF). Amount of tryptophan produced and activities of tryptophan synthase of $trpR^{-}$ mutant (KC 100) and $tyrR^{-}$ mutnat(KC 105) containing recombinant plasmid pSC 101-trp operon were increased by 30-40% as compared with KC 99(pheA tyrA trpE) containing recombinant plasmid pSC 101-trp operon. Activities of tryptophan synthase and production of tryptophan of KC 108 ($His^{-}$) and KC 109($Arg^{-}$) containing recombinant plasmid pSC 101-trp operon were increase by two-fold as compared with KC 107 containing pSC 101-trp operon.

• Korean Journal of Microbiology
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• v.32 no.2
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• pp.115-119
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• 1994

Replication control region of pSBK203, a chloramphenicol acetyltransferase conferring plasmid from Staphylococus aureus was cloned and its nucleotide sequence has been determined. Base sequence homology of this copy control region with those of plasmids belonging to pT181 family was obtained and analyzed. Copy number of four copy mutants derived by addtion or deletion of nucleotides in unique XbaI recognition site in copy control region of pSBK203 was also determined.

• Korean Journal of Microbiology
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• v.29 no.4
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• pp.226-229
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• 1991

In our laboratory a R plasmid pKU10 was isolated from Pseudomonas and its characteristics were investigated. In this study, as a basic work to improve its utility as a cloning vehicle, restriction patterns of pKU10 were analyzed for other various restriction enzymes in addition to restriction evdonucleases previously examined. As a result, pKU10 DNA has two cleavage sites for ClaI and HpaI, and three sites for AvaI. The restriction map of pKU10 was supplemented with AvaI, ClaI, and HpaI. From the result of this experiment, the usefulness of PKU10 as a cloning vector in Pseudomonas will be enhanced by constructions of mini-plasmid or hybrid plasmids.