• Korean Journal of Microbiology
• /
• v.25 no.3
• /
• pp.165-172
• /
• 1987

An expression vector in a mammalian cell was constructed using the origin of replication (OR) and the promoters of SV40. The plasmid pSVOE was constructed by inserting SV40 DNA fragment (1, 118bp) containing SV40 OR and promoters into pBR322-1, and then a multiple cloning sequence was inserted at the immediate downstream of the late promoter of SV40 in the pSVOE vector. The plasmid was named pSVML. As a selection marker, thymidine kinase gene of herpes simplex virus with its promoter was inserted into EcoRI site of pSVML and the recombinant was named pSVML-TKp. To test the expression capacity of foreigen gene inserted at the multiple cloning site of pSVML, the thymidine kinase gene without its own promoter was inserted at the BamHI site of pSVML. The recombinant was named pSVML-TK. These plasmids, pSVML-TKp and pSVML-TK, were transfected into COS cells with calcium phosphate precipitation method. The thymidine kinase activity was significantly increased in both transfected cells.

• Korean Journal of Microbiology
• /
• v.22 no.3
• /
• pp.183-189
• /
• 1984

In order to find specific acting site of Rec A protein in plasmic polymerization in E. coli, we randomly deleted various part of pEC-3 (a derivative of pBR322) with SI nuclease treatment. Self-ligated plasmids were introduced into E. coli WA802(Rec $A^+$). A number of colonies were analyzed if they contained monomeric or polymeric plasmids by gel electrophoresis. The plasmid (pEC-43), which was deleted the region of tetracycline gene, revealed only monomeric form in Rec $A^+$ E. coli. When two plasmids, pEC-3 and pEC-43, were co-transformed in the same E. coli, the original pEC-3 showed polymerization but pEC-43 revealed monomeric form only. These results suggest that Rec A protein requires the specific site for polymerization.

• Korean Journal of Microbiology
• /
• v.50 no.1
• /
• pp.22-26
• /
• 2014

Protocatechualdehyde (PA) is phenolic compound having antioxidative and antitumor activities. PA was purified from supernatant of Streptomyces lincolnensis M-20. In the presence of copper ion, PA acted as pro-oxidant. The antioxidant activity was assessed with the 1,1-diphenyl-2-picrylhydrazyl (DPPH) assay, and the pro-oxidant effect of PA on DNA damage as pBR322 plasmid DNA-cleaving agents in the presence of Cu(II) ions was investigated. The involvement of reactive oxygen species (ROS) in the DNA damage was confirmed by the inhibition of the DNA breakage by using glutathione (GSH), specific scavenger of ROS. When the increase in ROS reaches a certain level (the toxic threshold), it may trigger cell death. The formation of the PA/Cu(II) chelate complex was confirmed by reaction with ethylenediamine-tetraacetic acid (EDTA), a well-known chelating agent for metal ions, by using UV/Vis spectroscopic analysis.

• Microbiology and Biotechnology Letters
• /
• v.17 no.2
• /
• pp.154-159
• /
• 1989

Chromosomal DNA fragments of Bacillus sp. YA-14 isolated from soil as a potent xylan hydrolyzing bacterium, were ligated to a vector plasmid, pBR322, and used to transfer Escherichia coli HB101 cells. The recombinant plasmid pYDC21 was found to enable the transformants to produce xylanase. pYDC21 was found to contain the 3 kb HindIII fragment originated from the Bacillus sp. YA-14 chromosomal DNA by southern hybridization. The optimum temperature and pH for the reaction of xylanse produced by E. coli (pYDC21) were appeared at 50$_7$C and pH 7.0, respectiveiy. the xylanase enzyme was stable between pH 5.0 and 7.0 and maintained stably up to 4$0^{\circ}C$.

• Journal of Microbiology and Biotechnology
• /
• v.1 no.4
• /
• pp.251-255
• /
• 1991

A 16 kilobase (kb) HindIII fragment of alkalophilic Bacillus sp. YC-335 containing a gene for xylanase synthesis was inserted at the HindIII site of pBR322 and cloned in Escherichia coli HB101. After subcloning of recombinant plasmid pYS52, the 1.5 kb fragment was found to code for xylanase activity, and the hybrid plasmid was named pYS55. The DNA insert of the plasmid was subjected to restriction enzyme mapping, which showed that pYS55 had single site for PuvII and SstI in the 1.5 kb insert fragment. Southern hybridization analysis revealed that the cloned gene was hybridized with chromosomal DNA from alkalophilic Bacillus sp. YC-335. About 64% of the enzyme activity was observed in the extracellular and periplasmic space of E. coli HB10l carrying pYS55.

• Journal of Microbiology
• /
• v.34 no.1
• /
• pp.74-81
• /
• 1996

Two secretion vectors, pUBA240 and pUB340 were constructed by using the promoter and secretory signal region of the .alpha.-amylase gene from an .alpha.-amylase hyperproducing strain, Bacillus subtilis NA64. In this secretion vector system, various restriction enzyme sites are located immediately after the proregion of the .alpha.-amylase gene for easy replacement of various foregn structural genes. To evaluate this secretion vectors, the .betha.-lactamase gene of pBR322 was used as a reporter gene. The expressed and biologically active .betha.-lactamase was secreted into the culture broth from B. subtilis LKS86 transformants harboring each .betha.-lactamase secreting plasmid, pUBAbla and pUBSble. In both cases, more than 92% of expressed .betha.- lactamases were located idn the culture medium. The amount of the secreted .betha.-lactamase was about 80% of the total secreted proteins in the culture medium.

• Journal of Microbiology and Biotechnology
• /
• v.1 no.4
• /
• pp.227-231
• /
• 1991

A gene coding for a $\beta$-galactosidase of Bacillus subtilis HP-4 was cloned in E. coli JM109 by inserting HindIII digested fragment of B. subtilis HP-4 chromosomal DNA into the site of pBR322 and selecting recombinant transformant showing blue color on X-gal plate. The recombinant plasmid, named pBG109, was found to contain the 1.4 Kbp HindIII fragment originated from B. subtilis HP-4 chromosomal DNA by Southern hybridization. The cloned gene was stably maintained and expressed in E. coli JM109 and the pBG109 encoded $\beta$-galactosidase had the same enzymatic properties as those of $\beta$-galactosidase produced by B. subtilis HP-4.

• Journal of Microbiology and Biotechnology
• /
• v.2 no.3
• /
• pp.161-165
• /
• 1992

The gene encoding the bacteriolytic enzyme cell wall peptidoglycan hydrolase from alkalophilic Bacillus sp. was cloned in E. coli using pBR322 as a vector. A recombinant plasmid, designated pYTR451, was isolated and the size of the cloned HindIII fragment was found to be 4.8 Kb. The cell wall hydrolysis activity of an extract of the E. coli harboring the recombinant plasmid pYTR 451 was detected by SDS- polyacrylamide gel containing 0.2% (w/v) purified cell wall of Bacillus sp. The molecular weight of the enzyme was estimated to be about 27, 000 corresponding to the molecular weight of the Bacillus sp. bacteriolytic enzyme. The recombinant plasmid was found to contain the fragment originated from Bacillus sp. YJ-451 chromosomal DNA by Southern hybridization.

• Microbiology and Biotechnology Letters
• /
• v.22 no.6
• /
• pp.599-606
• /
• 1994

Bacillus stearothermomophilus, a strong xylan degrader, was confirmed to express multiple esterase activities in addition to the major xylanolytic enzymes. One of the genes encoding the esterases was isolated from the genomic library of B. stearothermophilus constructed with EcoRl restriction endonuclease and pBR322 plasmid. Three recombinant plasmids showing the tributyrin degrading activity were selected from approximately 7, 000 E. coli HB101 transformants, and were found to have the same insert of a 3.2 kb DNA fragment. Restriction mapping and hybridization studies revealed that the gene(estII) on the hybrid plasmid (pKMG7) had originated from the B. stearothermophilus chromosome, and was distinct from the estl, another esterase gene of B. stearothermophilus isolated in the previous work. The E. coli cells harboring pKMG7 produced an acetylxylan esterase that exibited similar substrate specificity to the esterase encoded by the estI gene.

• Journal of the Korean Chemical Society
• /
• v.42 no.3
• /
• pp.315-322
• /
• 1998

A series of new dimesogenic compounds were prepared and their thermotropic and liquid crystalline properties were studied by differential scanning calorimetry and on a hot-stage of a polarizing microscope. These compounds, 1,10-bis[2,5-bis(4-substitutedphenoxycarbonyl)phenoxy]decanes, consist of two bis(ρ-substitutedphenoxy)terephthalate units interconnected through a oxydecamethyleneoxy spacer on the central terephthaloyl units resulting in the structure of "H-shaped" dimeric twin compounds. The terminal substitutent groups were changed; X=-F, -H, -I, -Cl, -Br, $-NO_2,\;-CF_3,\;-OC_4H_9,\;-CN\;and\;-C_6H_5.$ The compounds with X=$-OC_4H_9,\;-CN\;and\;-C_6H_5$ were monotropically nematic. In contrast, the compounds with $X=-F, -H, -I, -Cl, -Br, $-NO_2\;and\;-CF_3$ were non-liquid crystals. The nematic group efficiency of these compounds was in the order of -C_6H_5>-CN>-OC_4H_9.$