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REFERENCE LINKING PLATFORM OF KOREA S&T JOURNALS
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The Korean Journal of Microbiology
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Journal DOI :
The Microbiological Society of Korea
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Volume & Issues
Volume 39, Issue 3 - Sep 2003
Volume 39, Issue 2 - Jun 2003
Volume 39, Issue 1 - Mar 2003
Volume 39, Issue 4 - Jan 2003
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Sequence and phylogenetic analysis of the phnS gene encoding 2-hydroxychromene-2-carboxylate isomerase in Sphingomonas chungbukensis DJ77
The Korean Journal of Microbiology, volume 39, issue 3, 2003, Pages 123~127
Sphingomonas chungbukensis DJ77 is able to metabolize phenanthrene as the sole carbon and energy source. The plasmid pUPX5 includes phnS gene encoding 2-hydroxychromene-2-carboxylate (HCCA) isomerase, which is needed for phenanthrene and naphthanene degradation. We determined the nucleotide sequence of DNA fragment of 3271 bp which included the phnS gene. The fragment included an open reading frame of 594 bp which has ATG initiation codon and TAA termination codon and GGAA ribosomal binding site. The predicted amino acid sequence of the enzyme consists of 198 amino acids. The deduced amino acid sequence of the phnS enzyme exhibited 94％ identity with that of the corresponding enzyme in Sphingomonas aromaticivorans F199. The phnS gene is located downstream and in the same operon as phnQ and phnR, encoding a 2,3-dihydroxybiphenyl 1,2-dioxygenase and a ferredoxin component of biphenyl dioxygenase, respectively.
Effect of GC Content on Target Hook Required for Gene Isolation by Transformation-Associated Recombination Cloning
The Korean Journal of Microbiology, volume 39, issue 3, 2003, Pages 128~134
Transformation-associated recombination (TAR) cloning is based on co-penetration into yeast spheroplasts of genomic DNA along with TAR vector DNA that contains 5'- and 3'-sequences (hooks) specific for a gene of interest, followed by recombination between the vector and the human genomic DNA to establish a circular YAC. Typically, the frequency of recombinant insert capture is 0.01-1% for single-copy genes by TAR cloning. To further refine the TAR cloning technology, we determined the effect of GC content on target hooks required for gene isolation utilizing the
mouse transgene as the targeted region. For this purpose, a set of vectors containing a B1 repeated hook and Tg AC-specific hooks of variable GC content (from 18 to 45%) was constructed and checked for efficiency of transgene isolation by radial TAR cloning. Efficiency of cloning decreased approximately 2-fold when the TAR vector contained a hook with a GC content ～
% versus ～40%. Thus, the optimal GC content of hook sequences required for gene isolation by TAR is approximately 40%. We also analyzed how the distribution of high GC content (65%) within the hook affects gene capture, but no dramatic differences for gene capturing were observed.
Expression of Secretion-dedicated Srb Homologue and Antifungal Activity of Bacillus lentimorbus WJ5
The Korean Journal of Microbiology, volume 39, issue 3, 2003, Pages 135~140
Bacillus sp. secretes high levels of extracellular enzymes into the culture medium. The signal recognition particle (SRP) and the SRP receptor play a central role in targeting pre secretory proteins to the translocase. By the analysis of the DNA microarray of B. lentimorbus WJ5, it was detected that WJ5m12, antifungal activity deficient mutant induced by gamma radiation, had a down-regulated expression of the SRP receptor gene (B. subtitis srb homologue, srbL). To determine the relationship of SRP receptor to antifungal activity, srbL of B. lentimorbus WJ5 was amplified by PCR and ligated into pQE30 vector, and then transferred into WJ5m12. The transformant, WJ5m12::srbL, recovered the antifungal activity. From the 2-DE analysis, the several presecretory proteins accumulated in the mutant cell and decreased to a level of the wild type in WJ5m12::srbL. It seems that the srbL could play an important role in the secretion of the antifungal activity related proteins of B. lentimorbus WJ5.
DNA Microarray Analysis of Gene Expression in Antifungal Bacterium of Bacillus lentimorbus WJ5
The Korean Journal of Microbiology, volume 39, issue 3, 2003, Pages 141~147
The simultaneous expression levels of antifungal activity related genes was analyzed by DNA microarray. We constructed DNA chips contained 2,000 randomly digested genome spots of the antifungal bacterium of Bacillus lentimorbus WJ5 and compared its quantitative aspect with 7 antifungal activity deficient mutants induced by gamma radiation (
). From the analysis of microarray hybridization by the Gene Cluster (Michael Eisen, Stanford Univ.), totally 408 genes were expressed and 20 genes among them were significantly suppressed in mutants. pbuX (xanthine permease, K222), ywbA (phosphotransferase system enzyme II, K393), ptsG (PTS glucose specific enzyme II ABC component, K877), yufO (ABC transporter (ATP-binding protein), K130l), and ftsY (signal recognition particle (docking protein), K868) were simultaneously down-regulated in all mutants. It suggested that they were supposed to be related to the antifungal activity of B. lentimorbus WJ5.
Purification and Assay of Extracellular Autolysin from Moraxella sp. CK-l
The Korean Journal of Microbiology, volume 39, issue 3, 2003, Pages 148~154
Moraxella sp. CK-l is known to inhibits the growth of Anabaena cylindrica, a cyanobacterium. It has been documented that the ability of this growth inhibition of Anabaena cylindrica was attributed to extracellular autolysin from Moraxella sp. CK-l. However, it remains to be elucidated identification and characterization of autolysin have yet been elucidated. In this study, we tried to purify and identify autolysin secreted from Moraxella sp. CK-l. Cells were grown in a complex liquid medium (BGC-11) and culture supernatants were collected, followed by ammonium sulfate fractionation. Fractions were further separated with anion exchange column, Mono-Q, in FPLC system and analyzed by SDS/PAGE. The fraction containing high autolysin activity showed a single distinct protein peak in anion column and molecular mass of about 17 kDa in SDS/PAGE. Nterminal amino acid sequencing of the protein was analyzed, of which result showed the homology with some proteases, including extracellular serine protease, Dichelobacter nodosus.
Changes in Lipids- and Fatty Acids Compositions in Response to Growth Temperature of Streptomyces viridochromogenes
The Korean Journal of Microbiology, volume 39, issue 3, 2003, Pages 155~160
The wild type and two morphological variants of Streptomyces viridochromogenes were studied for their lipidand fatty acid compositions at different incubation temperatures. It showed that a decrease in triacylglycerol content was closely linked to the aerial mycelium formation. Phospholipids showed no characteristic changes, except that the contents of phosphatidylethanolamine were clearly high for aerial mycelium deficient strain BR2 grown at
. The strain BR2 also presented unidentified aminolipids with various
values. Among the aminolipids, ornithinolipid increased gradually during the cultivation for all strains. The changes in fatty acid compositions showed a temperature dependency that the proportion of unsaturated acids decreased as the growth temperature increased. The proportion of straight chain saturated fatty acids decreased as the aerial mycelium developed, and it was most evident for the mutant strain M13 with more extensive aerial mycelium. The mutant strain BR2 presented significantly higher level of iso branched odd numbered saturated fatty acids.
A Study on Antibacterial Activity and Seroprevalence of Ornithobacterium rhinotracheale Isolated from the Domestic Chickens
The Korean Journal of Microbiology, volume 39, issue 3, 2003, Pages 161~165
Ornithobacterium rhinotracheale (OR) is a recently described gram-negative rod-shaped bacterium associated with respiratory tract infection in poultry. In order to investigate current occurrence of OR infection and to evaluate antibiotic susceptibility, the prevalence of OR antibody in domestic chickens were examined and the minimal inhibitory concentrations(MICs) of 8 antibiotics for 11 OR isolates was determined. All isolates tested were mostly susceptible to three antibiotics, ampicillin (MICs ranging from 0.38
/ml to 2
/ml), tetracycline (MICs 0.094～3
/ml) and doxycycline (MICs 0.047～4
/ml) but resistant to genatmicin. Ciprofloxacin, norfloxacin, enrofloxacin, and ofloxacin gave most isolates inhibition only in case of a higher concentration (MICs ranged in most cases from 3
/ml to 48
/ml). Out of 188 chicken flocks including broilers, broiler breeders, and layers, seropositive flock to OR were detected in 5 broilers (4%), 17 broiler breeders (50%), and 16 layers (55.2%), using commercial OR enzyme-linked immunosorbent assay(ELISA) kits. It suggested that OR infection was widespreaded in poultry farms in Korea.
Primer Evaluation for the Detection of Toxigenic Microcystis by PCR
The Korean Journal of Microbiology, volume 39, issue 3, 2003, Pages 166~174
Microcystin produced by cyanobacteria in surface waters, such as eutrophic lake and river, is a kind of serious environmental problems due to its toxicity to human and wild animals. Microcystin is synthesized nonribosomally by the large modular multi-functional enzyme complex known as microcystin synthetase encoded by the mcy gene cluster. Amplification of mcy genes by PCR from cultures and environmental samples is a simple and efficient method to detect the toxigenic Microcystis. In order to evaluate primers designed to detect toxic microcystin-producing strains, 17 cyanobacterial strains and 20 environmental samples were examined by PCR with 7 pairs of primers. Some microcystin-producing cyanobacteria were not detected with FAA-RAA, TOX4F-TOX4R and FP-RP primers. The fragment of unexpected size was amplified with NSZW2-NSZW1 primers in Microcystis strains isolated from the lakes in Korea. TOX1P-TOX1F primers failed in amplification of toxin-producing strains. Only MSF-MSR and TOX2P- TOX2F primers amplified the fragments of mcy genes from 11 strains of microcystin-producing Microcystis. The water samples taken from 20 lakes in Korea were analyzed by PCR using each of the primers. In all the water samples, cyanobacteria capable of producing microcystin were detected by the PCR with TOX2P-TOX2F primers. These results indicate that TOX2P-TOX2F primers are better than the other primers for detection of microcystin-producing Microcystis strains in Korea. The nucleotide sequences of mcy gene in Microcystis aeruginosa NIER10010 suggest genetic diversity of Korean isolates.
Utilization of Various Electron Acceptors in Shewanella putrefaciens DK-l
The Korean Journal of Microbiology, volume 39, issue 3, 2003, Pages 175~180
Microbial Fe(III) reduction is an important factor for biogeochemical cycle in anaerobic environments, especially sediment of freshwater such as lakes, ponds and rivers. In addition, the Fe(III) reduction serves as a model for potential mechanisms for the oxidation of organic compounds and the reduction of toxic heavy metals, such as chrome or uranium. Shewanella putrefaciens DK-1 was a gram-negative, facultative anaerobic Fe(III) reducer and used ferric ion as a terminal electron acceptor for the oxidation of organic compounds to
or other oxidized metabolites. The ability of reducing activity and utilization of various electron acceptors and donors for S. putrefaciens DK-1 were investigated. S. putrefaciens DK-1 was capable of using a wide variety of electron acceptor, including
, Fe(III), AQDS, and Mn(IV). However, its ability to utilize electron donors was limited. Lactate and formate were used as electron donors but acetate and toluene were not used. Fe(III) reduction of S. putrefaciens DK-l was inhibited by the presence of either
. Further S. putrefaciens DK-1 used humic acid as an electron acceptor and humic acid was re-oxidized by nitrate. Environmental samples showing the Fe(III)-reducing activity were used to investigate effects of the limiting factors such as carbon, nitrogen and phosphorus on the Fe(III) reducing bacteria. The highest Fe (III) reducing activity was measured, when lactate as a carbon source and S. putrefaciens DK-1 as an Fe(III) reducer added in untreated sediment samples of Cheon-ho and Dae-ho reservoirs.
The Detection and a Quantitative Evaluation of Viable but Non-Culturable Soil Bacteria Using a Modified Direct Viable Count Method
The Korean Journal of Microbiology, volume 39, issue 3, 2003, Pages 181~186
This study was performed to analyze quantitatively the number of living bacteria in forest soil samples collected from Mt. Keryong using improved direct viable count (DVC) and plate count (PC) methods. The number of living bacteria by DVC comprised 18～44% of the total direct count (TDC), whereas the number of living bacteria by PC was less than 1% of TDC. These results showed that viable but non-culturable (VBNC) bacteria existed in the soil with high percentages. Besides, DVC was proved to make it possible to make a quantitative detection of the VBNC bacteria. On the other hand, upon measuring the value from the conventional nutrient broth (NB) and
folded diluted nutrient broth (DNB), the values from the DNB showed 5 to 10 times higher than those from the conventional NB medium. These results indicate that oligotrophic bacterial groups, which could multiply in the low nutrient broth, abundantly exist in the soil ecosystem. It would also be possible to apply this kind of method to other substrate to make a quantitative detection of soil bacterial groups.
Isolation of Myxobacteria from Soil and RFLP Analysis of 16S rDNA Fragments.
The Korean Journal of Microbiology, volume 39, issue 3, 2003, Pages 187~191
In an attempt to isolate myxobacteria from soil samples, we isolated swarm and fruiting body forming bacteria that have bacteriolytic activity on Coli-spot agar plate. For the classification of myxobacteria, 16S rDNA RFLP patterns were analyzed. Amplified 16S rDNAs of myxobacteria type strains (Family I, II, III and IV), negative control strains and soil-isolates were restricted with HaeIII, EcoRI and EcoRV, respectively. We found that the soil-isolates belongs to myxobacteria Family I, II, III.
Isolation of Lactobacillus plantarum HB1 from Tongchimi and Its Nitrite-Scavenging Effect
The Korean Journal of Microbiology, volume 39, issue 3, 2003, Pages 192~196
To obtain large pools of lactic acid bacteria, a strain was isolated from Tongchimi. Through its sugar fermentation and analysis of 16S rRNA gene, it was identified to be Lactobacillus plantarum HB1. This strain is Gram-positive and catalase-negative. In the range of 1～88 bp in the HB1 16S rRNA gene, the HB1 strain was homologous with other L. plantarum strains by almost 100%, and in the range of the rest 32 bp, the HB1 strain showed considerable variation, compared to other strains. Nitrate which may exist in radish can be easily converted to nitrite. The nitrite interacts with amine, and becomes nitrosamine which may cause stomach cancer. The culture obtained by HB1 strain could eliminate 400
nitrite within 1.5 hr. It is necessary to isolate specific components which are involved in nitrite elimination in the culture and to study on its mechanism.
In Situ PCR on the Glass Slide Using the Conventional DNA Thermal Cycler
The Korean Journal of Microbiology, volume 39, issue 3, 2003, Pages 197~200
In order to establish effective in situ PCR on the glass slide using the conventional DNA thermal cycler, several parameters should be considered. These include full accessibility of PCR reagents into the cells, prevention of diffusing PCR products out of the cells, loss of PCR reagents by nonspecific adherence onto the glass slide, dryness of PCR reagents by heat, and heat conductivity from the heat block to the glass slide. Especially, to guarantee the full accessibility of PCR reagents to sample, relatively higher concentration of PCR reagents (particularly 4.5 mM of
) was required while 5 to 10 units/50
reaction of Taq enzyme was enough as long as the step of pre-PCR incubation was included. Dryness of sample was prevented by addition of distilled water into the empty slots in the heat block, thereby providing the reproducible temperature-time profile of PCR. Observed temperature was lower than the programmed temperature by 3 to
Partial Cloning of Genes for Lignin Degrading Enzymes in Trametes versicolor
The Korean Journal of Microbiology, volume 39, issue 3, 2003, Pages 201~205
Laccase, lignin- and manganese peroxidase are implicated in the lignin degradation. The nucleotide sequences of four copper-binding domains in fungal laccases, and heme-binding domains of lignin- and manganese peroxidases are well conserved, and therefore these short fragments can be used for the PCR for the gene amplification. We synthesized several PCR primers according to their sequences, and run PCR to amplifiy the lignin degrading genes of Trametes versicolor isolated in Korea. PCR products were cloned with pGEM-T vector in order to determine their nucleotide sequences. A laccase fragment (1.3 kb) showed 65-97％ homologies, lignin peroxidase fragment (185 bp) showed 80-95％ homologies, and manganese peroxidase fragment (443 bp) showed 61-83％ homologies when compared with other white-rot fungal enzymes.