• Journal of Periodontal and Implant Science
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• v.25 no.3
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• pp.679-693
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• 1995

The present study has been performed to see the intrafamilial transmission of periodontopathic organism Actinobacillus actinomycetemcomitans in Koreans having various froms of periodontal diseases. 17 clinical isolates from 8 periodontal patients and 20m clinical isolates from their 8 family members were grown anaerobically for the serotyping and the extraction of genomic DNA. The DNA was digested with restriction endonucleases (EcoRI+HindIII) and plasmid pAA 2097(kindly provided by Dr. DiRienzo, Univ. of Pennsylvania) including 4.7kb-size randlomly clone probe for restriction length pleomorphism analysis(RFLP). RFLP patterns of reference serotypes a, b, c, d, and e were used as the genotypes A, B, C, D, and E, respectively for comparison of genotypes of clinical isolates. 28 out of total 37 clinical isoltes belonged to either one of 5 basic gentotypes and 9 remaining isolats did not fall into any types, and hence were designate as non-type(NT). Genotype C were the most frequently found one(35.1%) and genotype B has not isolated. Intrafamilial transmission of bacteria between spouses, brothers and sisters, and parents and their offsprings, resepctively could well be demonstrated by comparing RFLP patterns. There were not any specific genotypes which showed predominance over others in terms of transmission.

• Microbiology and Biotechnology Letters
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• v.20 no.3
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• pp.263-270
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• 1992

For the selection of powerful antagonistic bacterium for biological control of soil borne Eminia carotovora subsp. carotovora causing rot of vegetable, excellent strains (S4, S14, 565) were selected from 1,196 strains of bacteria which were isolated from rhizosphere in vegetable root rot-suppresive soil. Strains were identified to be Pseudomonas species with Api 20NE kit. Antagonistic substance was produced in 523 synthetic broth medium at pH 7~8 and $30^{\circ}C$ during 3 days culture. The substance was stable in the pH range of 6 to 9. When the basal medium was supplemented with mannitol and sorbitol as carbon source and calcium chloride as metal salt, the production of the inhibitory substance was increased. The inhibitory acitivity was increased by the addition of fertilizer in soil. The isolated strains were resistant to the agricultural chemical such as benomyl and fosethyl-Al-folpet, and the antibiotics such as penicillin and lincomycin. We had found that Pseudomonas sp. S14 strain had a single plasmid. After treated with acridin orange for curing, we confirmed the existence of antagonistic gene in the chromosomal DNA.

• Parasites, Hosts and Diseases
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• v.44 no.4 s.140
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• pp.303-312
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• 2006

Interactions between GRA proteins of dense granules in Toxoplasma gondii and host cell proteins were analyzed by yeast two-hybrid technique. The cMyc-GRA fusion proteins expressed from pGBKT7 plasmid in Y187 yeast were bound to host cell proteins from pGADT7-Rec-HeLa cDNA library transformed to AH109 yeast by mating method. By the selection procedures, a total of 939 colonies of the SD/-AHLT culture, 348 colonies of the $X-\alpha-gal$ positive and PCR, 157 colonies of the $X-\beta-gal$ assay were chosen for sequencing the cDNA and finally 90 colonies containing ORF were selected to analyze the interactions. GRA proteins interacted with a variety of host cell proteins such as enzymes, structural and functional proteins of organellar proteins of broad spectrum. Several specific bindings of each GRA protein to host proteins were discussed presumptively the role of GRA proteins after secreting into the parasitophorous vacuoles (PV) and the PV membrane in the parasitism of this parasite.

• The Journal of Korean Society of Virology
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• v.26 no.1
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• pp.77-90
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• 1996

Nucleocapsid protein (NP)which exists in the particle of hantavirus and surrounds the viral RNA genome is one of the major structural proteins and plays role of antigen to elicit the antibody detected predorminantly right after infection of the virus in the patients of hemorragic fever with renal syndrome (HFRS)or experimental animals. NP is important target antigen in serological diagnostic system of HFRS utilizing whole antigens from the native virus particle, such as IFA, ELISA and Western blotting. Therefore, the preparation of this protein in the level of higher quantity and purity is desirasble for developed dianosis of the disease. The purpose of this study is the cloning of NP gene which exists in the S genome segment of Maaji (MAA) virus and expression of the gene to obtain qualified, genetically engineered NP to be utilized as an immunodiagnostic antigen. First of all, for the purpose of amplifing the MAA-NP gene by PCR, the specific primers were built from the known nucleotide sequence of Hantaan viral NP gene. The viral cDNA of the NP gene was synthesized by using the primers and RNase $H^-$ AMV reverse transcriptase. Thereafter, using this cDNA as a template, the NP gene was amplified specifically by Taq DNA polymrerase. The pT7blue (R)T-overhang vector systems were used for cloning of the amplified NP gene. The expression system was consisted of BL21 (DE3)pLysS and pET16b as a host and a plasmid repectively. Into Ndel site of pET16b, NP gene was ligated with cohesive end for the expression. Insertion of NP gene in the plasmid was confirmed by PCR and mini prep methods. For expression, IPTG was used and the expressed protein was characterized by Western blotting. The MAA-NP was expressed as the form of inclusion body (insoluble fraction)and the protein purified by affinity and metal chealating columns reacted specifically with the sera from patients of HFRS as to be tested by ELISA and Western blotting.

• Journal of Microbiology and Biotechnology
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• v.3 no.1
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• pp.31-38
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• 1993

A bacterial strain KY-l isolated from sewage was able to produce an extracellular agarase(agarose 4-glycanohydrolase. EC 3.2.1.81). The strain KY-1 was identified as Cytophaga fermentans subsp. agarovorans based on its morphological and physiological characteristics. The agarase was purified by ammonium sulfate precipitation followed by DEAE-Sephadex A-50. Bio-Gel P-100. and CM-Cellulose column chromatography. The molecular weight of the purified enzyme was 24 kDa by SDS-polyacrylamide gel electrophoresis. The optimum temperature and pH for the enzyme activity were 30^{circ}C and 7.5, respectively. The enzyme activity was significantly inhibited in the presence of 0.1 mM $HgCl_2$. whereas it was elevated 3 times by $MnSO_4$ at 1 mM concentration. The Km value and Vmax were 16.67 mg/ml and 3.77 unit/ml.min. The agarase gene was cloned into Escherichia coli MC1061 using the plasmid vector pBR322. A 1.4 Kb DNA fragment of PstI-digested chromosomal DNA of C. fermentans KY-l was inserted into the PstI site of pBR322. expressed in the E. coli. and up to 60% of the total enzyme was extracellularly secreted. Enzymatic properties of the extracellular agarases produced by both the transformant and the donor were very similar in terms of optimal pH and temperature.

• Microbiology and Biotechnology Letters
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• v.26 no.1
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• pp.23-33
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• 1998

The entire sequence of a 4,214,810 bp genome of the Bacillus subtilis 168 has been determined by an international project, and the completion has been announced on July 19, 1997. For the sequencing project an international consortium was established and 25 European, 7 Japanese laboratories, 2 biotechnology companies, and our laboratory participated in the project. Within this framework we determined the complete nucleotide sequence of a 53,289 bp fragment upstream of the odhA gene (181 $^{\circ}$) of the B. subtilis 168 chromosome. On the basis of the published DNA sequences of the B. subtilis sspC and odhA genes, we obtained genomic fragments by plasmid rescue and long-range PCR. The sequenced fragment contains 56 putative open reading frames (designated yojA-yolI and 9 known genes (sspC, cge cluster, orfE5, orfRMl and odhA), in which we found many interesting features. In addition, the entire nucleotide sequence of a 53,289 bp region enabled us to revise the current genetic map of this region.

• Biomedical Science Letters
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• v.12 no.3
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• pp.139-143
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• 2006

Jopock (Jpk) has previously been ascertained that induces both bacterial and mammalian cell death. The Escherichia coli cells expressing Glutathion S-transferase (GST) fused Jpk showed elongated phenotype and inhibited cell growth which led eventual cell death. In an attempt to search the genetic suppressor of the lethal protein Jpk in bacterial cells, we constructed a unidirectional protein expression vector inserting tac promoter next to the C-terminus Jpk in pGEX-Jpk. The function of additional tac promoter was confirmed by substituting lac promoter in Plac-TOPO plasmid. The cells harboring plac- TOPO, which regulates $lacZ{\alpha}$ gene expression under lac promoter, formed blue colonies in 5-bromo-4-3 $indolyo-{\beta}-D-galactoside$ (X-gal) plate. When lac promoter was changed to tac promoter, same results were observed. Since the addition of tac promoter did not affect the toxic effect of Jpk, the pGEX-Jpk-ptac could be a useful vector for the screening of suppressor(s) for Jpk, in which GST-Jpk and a putative Jpk-suppressing protein are coexpressing from two unidirectional tac promoters, which response to the same inducer, $isopropyl-{\beta}-D-thiogalactopyranoside (IPTG)$.

• Applied Biological Chemistry
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• v.44 no.4
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• pp.251-256
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• 2001

To develop biofertilizer solubilizing inorganic phosphate, a bacterium having high abilities to solubilize inorganic phosphate were isolated from cultivated soils. The strain was identified to Aeromonas hydrophila DA57, based on the physiological and biochemical properties. The optimum temperature and initial pH to solubilize insoluvle phosphate in sucrose minimal medium were $30^{\circ}C$ and pH 7.0, respectively. In these conditions phosphate solubilizing activities of the strain against three types of insoluble phosphate were quantitatively determined. It was possivle to distinguish between solubilization through release of gluconic acid and still unknown mechanism. Aemmonas hydrophila DA57 harbored a 4.5 kb cryptic plasmid.

• Korean Journal of Microbiology
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• v.31 no.3
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• pp.179-183
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• 1993

A restricted facultatively methanol-oxidizing bacterium, Methylovorus sp. strain SS1, was isolate dfrom soil samples from Kuala Lumpur, Malaysia, through methanol-enrichment culture technique. The isolate was nonmotile Gram-negative rod and did not have complex internal membrane system. The colonies were small, pale-yellow, and raised convex with entire margin. The cell did not produce any spores and capsular materials. The cell was obligately aerobic and exhibited catalase, but no oxidase, activity. Plasmid, carotenoid pigment, and poly-.betha.-hydroxybutyric acid were not found. The guanine plus cytosine content of the DNA was 55%. The isolate was found to grow only on methanol methylamine, or glucose. Growth factors were not required. Cells growing on methanol was found to produce extracellular polysaccharides containing glucose, lactose, and fructose. Growth was optimal (t$_{d}$= 1.7) with 0.5%(v/v) methanol at 40.deg.C and pH 6.5. No Growth was observed at over 60.deg.C. Cell-free extracts of the methanol grown cells exhibited the phenazine methosulfate-linked methanol dehydrogenase activity Methanol was found to be assimilate dthrough the ribulose monophosphate pathway.y.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.29 no.5
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• pp.637-642
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• 1996

We isolated a marine bacterium, Pseudomonas sp,, which could produce the enzyme of alginate lyase, and cloned the alginate lyase gene in Escherichia coli. The cloned DNA was overexpressed with approximately $50\%$ amount of total proteins. In addition, the expressed proteins were not secreted into the medium, and most of them existed in the cytoplasm by the soluble form, but not observed any inclusion body by TIM. For the optimum enzyme activity, temperature was $20^{\circ}C$, pH was 7.0, and Km and Vmax values of the enzyme were $0.4\%$ and 625 units/mg, respectively.