• Journal of Microbiology
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• v.44 no.3
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• pp.255-262
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• 2006

Titanium and its alloys are technically superior and cost-effective materials, with a wide variety of aerospace, industrial, marine, and commercial applications. In this study, the effects of titanium ions on bacterial growth were evaluated. Six strains of bacteria known to be resistant to both metal ions and antibiotics were used in this study. Two strains, Escherichia coli ATCC 15489, and Pseudomonas aeruginosa ATCC 10145, proved to be resistant to titanium ions. Plasmid-cured p. aeruginosa resulted in the loss of one or move resistance markers, indicating plasmid-encoded resistance. The plasmid profile of p. aeruginosa revealed the presence of a 23-kb plasmid. The plasmid was isolated and transformed into $DH5{\alpha}$. Interestingly, the untransformed $DH5{\alpha}$ did not grow in 300 mg/l titanium ions, but the transformed $DH5{\alpha}$ grew quite well under such conditions. The survival rate of the transformed $DH5{\alpha}$ also increased more than 3-fold compared to that of untransformed $DH5{\alpha}$.

• Journal of Wetlands Research
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• v.13 no.3
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• pp.697-705
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• 2011

The aim of this study was to examine ozonation disinfection efficiency for Escherichia coli DH5alpha removal, containing the multi-resistance plasmid pB10 as well as chlorination disinfection efficiency. In addition, plasmid pB10 removal rates were estimated by ozonation and chlorination. The removal efficiency of pB10 via ozonation was about 2 to 4 times higher than chlorination. High removal efficiency of pB10 is likely due to OH radical produced during ozonation. These results suggest that integration of advanced oxidation process such as ozonation (or photocatalytic oxidation) with conventional disinfection such as chlorination may be needed for effective control of antibiotic resistant bacteria and genetic materials.

• Food Science and Preservation
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• v.20 no.2
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• pp.250-256
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• 2013

To verify the multiplication of microorganisms on the surface of strawberries, the fate of E. coli $DH5{\alpha}::gfp$ at different temperatures, times and strawberry extract concentrations were measured. The population of E. coli $DH5{\alpha}::gfp$ rapidly increased by 7.36~7.78 log CFU/g at $25{\sim}30^{\circ}C$ for 24 hr and slowly increased by 6.49~8.49 log CFU/g at $10{\sim}20^{\circ}C$ for 48 hr. However, E. coli $DH5{\alpha}::gfp$ did not grow at $10{\sim}15^{\circ}C$ on the surface of the strawberries, regardless of the contact times with the bacterial suspension. E. coli $DH5{\alpha}::gfp$ reached 1.52~3.26 log CFU/g at $20^{\circ}C$ as the contact frequency increased from two to six times. The contact frequencies did not significantly differ. In the case of the six-time contact on the surface of the strawberry at 25 and $30^{\circ}C$, the E. coli $DH5{\alpha}::gfp$ increased by 5.17 and 5.01 log CFU/g. The effects of the strawberry extracts on the growth of E. coli $DH5{\alpha}::gfp$ showed that sterilization and non-sterilization do not affect the growth of microorganisms for 96 hr. In the minimal broth, the growth of E. coli $DH5{\alpha}::gfp$ increased by 1 log CFU/g for 96 hr. In less than 50 percent of the strawberry extracts, the growth rate of E. coli $DH5{\alpha}::gfp$ was higher than in the control and increased by 4 and 5 log CFU/g at 50 and 25 percent of strawberry extracts, respectively. Therefore, E. coli $DH5{\alpha}::gfp$ can multiply and survive on the surface of strawberries when it comes into contact with the fruit extract.

• The Korean Journal of Food And Nutrition
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• v.23 no.4
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• pp.526-530
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• 2010

Isopentenyl diphosphate(IPP) isomerization to dimethylallyl diphosphate(DMAPP) is an important step for the efficient production of isoprenoids such as lycopene, ${\beta}$-carotene, astaxanthin, etc. The type II isopentenyl diphosphate isomerase gene from Synechocystis sp. PCC6803(sll1556, Syidi2) was cloned and expressed in Escherichia coli $DH5{\alpha}$. When E. coli $DH5{\alpha}$ harboring lycopene synthesis genes, crtE, crtB, and crtI and mevalonate pathway genes, MvK1, MvK2, and Mvd, was cultured on LB medium containing mevalonate, the strain grew very slowly be due to the toxicity of isopentenyl diphosphate derived from mevalonate. When Syidi2 was introduced to E. coli $DH5{\alpha}$ harboring the lycopene synthesis genes and mevalonate pathway genes, growth on mevalonate medium was fully restored and the colony showed red color indicating lycopene formation. The growth rate of the mutant strain, E. coli $DH5{\alpha}$(idi::${\Delta}km$), was very slow because of IPP accumulation and DMAPP deprivation. Ultimately the idi mutant was complemented by introducing the Syidi2 gene.

• KSBB Journal
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• v.17 no.6
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• pp.571-576
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• 2002

Recombinant E. coli DH5 ${\alpha}$/pSB311 was made by cloning the genes encoding bacterial luciferase and aldehyde substrate proteins from Photohabdus luminescense, to complement defects of Lumistox, which is normally used in bioassays to monitor toxic substances in water environmental systems. The conditions for stable light production by the recombinant strains were investigated with respect to cell growth stage, cell number, and buffer conditions. The optimum growth stage was a middle-exponential stage with an OD$_{660nm}$ value of 0.6-0.7. ADout 10$^{6}$-10$^{7}$ cells per test tube was optimum for stable light emission. The effect of buffer was not significant if an optimum viable cell number was maintained. The bioluminescence of the recombinant E. coli harboring the lux operon of Photohabdus luminescense was not affected by temperature, while the bioluminescence of Lumistox was temperature sensitive. The recombinant E. coli was more sensitive to heavy metals (Cd, Cu, Hg, Zn) than Lumistox, because it does not require high concentrations of NaCl in the buffer.

• Journal of Microbiology and Biotechnology
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• v.4 no.2
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• pp.102-107
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• 1994

The $\beta$-1,4-endoglucanase gene from Actinomyces sp. 40 was cloned into Escherichia coli DH5$\alpha$ with pUC19. Chromosomal DNA from Actinomyces sp. 40 was cleaved with the restriction enzyme Sau3AI and ligated into pUC19 for the transformation of Escherichia coli DH5$\alpha$. Positive clones of $\beta$-1,4-endoglucanase gene were detected as the clear zones on a medium supplemented with carboxymethylcellulose (CMC). This transformant possessed a single plasmid, designated pDS1, which contained the vector DNA and a 3.5 kilobase (kb) Sau3AI insertion fragment encoding endoglucanase. The size of the cloned fragment was reduced to 2.0 kb. The endoglucanase activity produced by the E. coli DH5$\alpha$ (pDS6) was higher than that of Actinomyces sp. 40 strain. The optimum pH and temperature of the cloned enzyme were pH 4.0$\sim$5.0 and 55$^{\circ}C$, respectively. The cloned enzyme was stable at 55$^{\circ}C$ or below and in buffer ranging from pH 4.0 to 7.0. The enzyme degraded CMC but did not degrade xylan, cellobiose, and methyl-umbelliferylcellobiopyranoside (MUC).

• Proceedings of the Microbiological Society of Korea Conference
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• 2000.10a
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• pp.128-135
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• 2000

The thiosulfate reductase gene (PhsABC) from Salmonella typhimurium was expressed in Escherichia coli in order to produce sulfide from inorganic thiosulfate and precipitate metals as metal sulfide complexes. A 5.1-kb DNA fragment containing the native phsABC and a 3.7-kb DNA fragment, excluding putative promoter and regulatory regions were inserted into expression vectors pTrc99A and pJB866, respectively. Upon expression of phsABC, E. coli DH5$\alpha$ harboring the phsABC constructs showed higher thiosulfate reductase activity and produced significantly more sulfide than the control strain (E. coli DH5$\alpha$) under both aerobic and anaerobic conditions. Among the four constructs, E. coli DH5$\alpha$ harboring pSB74 produced the highest level of thiosulfate reductase and removed most of heavy metals from solution under anaerobic conditions. In a mixture of 100 $\mu$M each of cadmium, lead, and zinc, the strain could remove $99\%$ of the total metals from solution within 10 hours. Cadmium was removed first, lead second, and zinc last. In contrast, a negative control did not produce any measurable sulfide and removed very little metals from solution. These results have important implications for removal of metals from wastewater contaminated with several metals.

• KSBB Journal
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• v.13 no.1
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• pp.96-100
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• 1998

Two promoters (trc and P$\rho$) were inserted in PHA operon derived from Alcaligenes eutrophus to obtain high chain molecules of polyhydroxybutyrate (PHB) in recombinant Escherichia coli. Newly designed PHA operon was used to control the gene expressions of hydroxybutyric CoA and PHA polymerization, separately. Plasmids containing new synthetic operon was transformed into E. coli DH5$\alpha$ and analyzed for PHB production. Without induction of the PHA biosynthetic operon, PHA synthase which has low activity might supply low concentration of initiator during the polymerization reaction, resulting very high molecular weight of polymer. An increase of PHB average molecular weight was observed with decreased IPTG (isopropyl $\beta$ -Dithiogalactosidase) concentration. When no IPTG was added to the culture of E. coli DH5$\alpha$ /$\rho$ SJS1 which contained two promoters in PHA operon, high chain polymer having an average molecular weight of $2.5{\times}10^7$ was achieved. Analysis of the enzyme activities of PHA biosynthetic enzymes suggests that PHA synthase, the enzyme responsible for polymerizing 3-hydroxybutyric CoA, controls the molecular weight of PHB produced in vivo.

• Journal of Microbiology and Biotechnology
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• v.23 no.12
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• pp.1726-1736
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• 2013

Biosynthesis of indole-3-acetic acid (IAA) via the indole-3-pyruvic acid pathway involves three kinds of enzymes; aminotransferase encoded by aspC, indole-3-pyruvic acid decarboxylase encoded by ipdC, and indole-3-acetic acid dehydrogenase encoded by iad1. The ipdC from Enterobacter cloacae ATCC 13047, aspC from Escherichia coli, and iad1 from Ustilago maydis were cloned and expressed under the control of the tac and sod promoters in E. coli. According to SDS-PAGE and enzyme activity, IpdC and Iad1 showed good expression under the control of $P_{tac}$, whereas AspC was efficiently expressed by $P_{sod}$ originating from Corynebacterium glutamicum. The activities of IpdC, AspC, and Iad1 from the crude extracts of recombinant E. coli Top 10 were 215.6, 5.7, and 272.1 nmol/min/mg-protein, respectively. The recombinant E. coli $DH5{\alpha}$ expressing IpdC, AspC, and Iad1 produced about 1.1 g/l of IAA and 0.13 g/l of tryptophol (TOL) after 48 h of cultivation in LB medium with 2 g/l tryptophan. To improve IAA production, a tnaA gene mediating indole formation from tryptophan was deleted. As a result, E. coli IAA68 with expression of the three genes produced 1.8 g/l of IAA, which is a 1.6-fold increase compared with wild-type $DH5{\alpha}$ harboring the same plasmids. Moreover, the complete conversion of tryptophan to IAA was achieved by E. coli IAA68. Finally, E. coli IAA68 produced 3.0 g/l of IAA after 24 h cultivation in LB medium supplemented with 4 g/l of tryptophan.

• Fisheries and Aquatic Sciences
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• v.6 no.1
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• pp.20-26
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• 2003

A bacterial strain was isolated from seawater to screen for phytase activities. A colony had the highest activity and was identified as an Escherichia coli strain. Using primers derived from E. coli acid phosphatase appA sequence, we cloned a 1,495 bp DNA fragment connected with the pGEM-T vector. It was over-expressed under lac promoter combined with its native promoter in E. coli $DH5\alpha$. The expression of the phytase gene occurred during late exponential growth and the intracellular phytase production was 16.9 units/ml. The yield of recombinant phytate was 412-fold higher than that of wild type E. coli K13.