• Journal of Microbiology and Biotechnology
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• v.22 no.4
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• pp.555-562
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• 2012

PCR was performed to analyze the ${\beta}$-lactamase genes carried by ampicillin-resistant Vibrio spp. strains isolated from marine environments in Korea between 2006 and 2009. All 36 strains tested showed negative results in PCR with the primers designed from the nucleotide sequences of various known ${\beta}$-lactamase genes. This prompted us to screen new ${\beta}$-lactamase genes. A novel ${\beta}$-lactamase gene was cloned from Vibrio alginolyticus KV3 isolated from the aquaculture water of Geoje Island of Korea. The determined nucleotide sequence (VAK-3 ${\beta}$-lactamase) revealed an open reading frame (ORF) of 852 bp, encoding a protein of 283 amino acids (aa), which displayed low homology to any other ${\beta}$-lactamase genes reported in public databases. The deduced 283 aa sequence of VAK-3, consisting of a 19 aa signal peptide and a 264 aa mature protein, contained highly conserved peptide segments specific to class A ${\beta}$-lactamases including the specific amino acid residues STFK (62-65), SDN (122-124), E (158), and RTG (226-228). Results from PCR performed with primers specific to the VAK-3 ${\beta}$-lactamase gene identified 3 of the 36 isolated strains as V. alginolyticus, Vibrio cholerae, and Photobacterium damselae subsp. damselae, indicating the utilization of various ${\beta}$-lactamase genes including unidentified ones in ampicillin-resistant Vibrio spp. strains from the marine environment. In a mating experiment, none of the isolates transfered the VAK-3 ${\beta}$-lactamase gene to the Escherichia coli recipient. This lack of mobility, and the presence of a chromosomal acyl-CoA flanking sequence upstream of the VAK-3 ${\beta}$-lactamase gene, led to the assumption that the location of this new ${\beta}$-lactamase gene was in the chromosome, rather than the mobile plasmid. Antibiotic susceptibility of VAK-3 ${\beta}$-lactamase was indicated by elevated levels of resistance to penicillins, but not to cephalosporins in the wild type and E. coli harboring recombinant plasmid pKV-3, compared with those of the host strain alone. Phylogenetic analysis showed that VAK-3 ${\beta}$-lactamase is a new and separate member of class A ${\beta}$-lactamases.

• Korean Journal of Animal Reproduction
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• v.21 no.2
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• pp.167-176
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• 1997

This study was carried out to evaluate the potential for preselection of transgenic embryos prior to transfer into recipient animals. In these experiments, I used a 3.2 kb transgene which contained the neomycin resistance gene (neo) and lac Z gene driven by the $\beta$ actin promoter (iacZ Ineo). Oocytes were aspirated from abattoir ovaries, matured in TCM-199 supplemented with 10% fetal bovine serum (FBS), 5 ${\mu}\textrm{g}$/ml LH, 0.5 ${\mu}\textrm{g}$/ml FSH, 100 unit/ml penicillin, and 100 ${\mu}\textrm{g}$/ml streptomycin for 22 to 24 hrs then inseminated with a sperm suspension of 1 X 10$^6$ sperm/ml containing 5 ${\mu}\textrm{g}$/ml of heparin. At 18-20 hrs after insemination, cumulus cells were removed by vortexing and pronuclei of centrifuged zygotes microinjected with the lacZ/neo construct (3 ng/$\mu$l). All cultures were carried out in CR1aa with transfected BRL monolayers containing 3 mg/ml BSA, 20 $\mu$/ml NEM amino acids and 40 $\mu$I/ml BME amino acids. To identify the appropriate concentration of G418 for selection, non-microinjected zygotes were cultured in the presence of 0, 50, 100 and 200 $\mu$g/ml of G418. After 8 days of culture in these treatments, 44/145 (30.3%), 13/150 (8. 7%), 1/151 (0.7%) and 0/134 (0.0%) devel-oped to the blastocyst stage in 0, 50, 100 and 200 $\mu$g/ml of G418, respectively. A total of 1,127 zygotes were microinjected and placed into culture (without G418) and subsequently 710 (63.0%) cleaved. At 48 hrs post-injection, embryos ($\geq$2-cell) were randomly assigned to control (medium alone) or G418 (100 ${\mu}\textrm{g}$/ml) treatments. A control culture of 740 non-microinjected embryos from the same replicates of embryos were also placed into control medium. After 8 days in culture, 54/343 (15.7%) and 22/367 (6.0 %) of the microinjected embryos developed to the blastocyst stage in control and G418 media, respectively. A total of 151/740 (27.2%) of the non-microinjected embryos placed in the control medium developed to the blastocyst stage. The blastocysts in the control treatment had a mean of 70.7 ${\pm}$ 4.7 cells of which 23.1 ${\pm}$ 2.6 (32.7%) stained for $\beta$-Gal activity. B1astocysts in the G418 treatment had a mean of 48.8${\pm}$7.5 cells of which 40.3 ${\pm}$ 4.1 (82.6%) stained for $\beta$-Gal ac tivity (P<0.05). In the control treatment 26 of 30 (87.0%) blastocysts had some cells with $\beta$-Gal activity while all of the blastocysts in the G418 treatment had some cell with $\beta$-Gal ac tivity (P<0.05). However, ICM colonies in either control or G418 treatments were not expressed any epiblast cell except of trophetoderm celIs. The $\beta$-actin promoter/lacZ gene may not be e expression or silence expression in epiblast cells These results clearly show an enrichment of blastocysts expressing the transgene in the majority of their cells after culture in the presence of G418. The exogeneous gene was not express a and silence in ICM colonies especiallly epiblast cells except of trophectederm cells. Even though the higher rate cell number expressed of exogeneous gene in the G418 treatments, a total cell number was decrease significantly (P<0.05) of which might be also drop of the establishment of ES like-cell colonies and production of transgenic animals. However, futher studies need to determine the viability of these selected embryos and the avability of production of transgenic offspring.

• Journal of Mushroom
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• v.18 no.3
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• pp.221-233
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• 2020

This study was carried out to compare the growth characteristics, biological activities, β-glucan contents, sugar contents, and amino acid contents of 14 strains of Ganoderma spp. Among the 14 strains of Ganoderma spp., KMCC02960 (G. meredithae) and KMCC02932 (G. tropicum) showed excellent growth characteristics such as those with respect to the size and yield of fruiting bodies. The highest 2,2-diphenyl-1-picrylhydrazyl radical scavenging activity was observed in KMCC02932 (G. tropicum). The nitrite scavenging activities of KMCC02824 (G. lucidum) and KMCC02852 (G. neo-japonicum) were higher than those of the other strains. The total polyphenol contents of the extracts from KMCC02824 (G. lucidum) and KMCC02852 (G. neo-japonicum) were higher than those of the other strains. KMCC03018 (G. lingzhi) showed the highest β-glucan content of 33.4%. In an analysis of the 4 types of monosaccharides, 2 types of disaccharides, and 4 types of sugar alcohols, only KMCC02996 (G. weberianum) and KMCC03018 (G. lingzhi) were commonly detected out of the 14 strains of Ganoderma spp. Eighteen amino acids, including eight essential amino acids, were identified: the highest total amino acids and total essential amino acids were found in KMCC02932 (G. tropicum), which had the highest levels of tyrosine and phenylalanine. Although the contents of amino acids differed by strain, cysteine, tyrosine, and phenylalanine were the most abundant amino acids in the analyzed extracts.

• Plant Resources
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• v.1 no.2
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• pp.113-120
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• 1998

Soybean is an important crop because its seed has very high protein relative to others. The quality of soy protein is limited by the concentration of the sulfur-containing amino acids in the amino acid profile. Among the supply of various forms of 0.4mM sulfur as S nutrition during seed fill. only 0.4mM L-methionine can inhibit ${\beta}$-subunit synthesis completely and produce the highest glycinin-containing seeds. Compared to 0.4mM sulfate control, seeds supplied by 0.4mM L-methionine have lower ${\alpha}$-, no ${\beta}$-subunit, and highly increased glycinin without altering total protein concentration. Supply of 0.2mM cystine (0.4mM S) did not affect the accumulative pattern of seed storage protein (SSP) subunits. In the supply of L-methionine, 0.2mM treatment showed higher glycinin in seeds but 0.05mM resulted in lower glycinin than tile sulfate control. The relative abundance of ${\alpha}^`$-subunit was not altered by any N or S nutrition. Under 5mM nitrogen, protein concentration was increased about 3-5% by substituting ammonia for nitrate during seed fill independent of nutrition. The increase resulted in the only increase of 7S protein, mainly ${\beta}$-subunit. Our data suggest that the regulatory system of SSP genes responds to the balance between N and S assimilates supplied from mother plant. and controls the di fferential synthesis of their subunits for the maximum protein accumulation in developing soybean seed.

• Journal of Microbiology and Biotechnology
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• v.18 no.1
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• pp.160-166
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• 2008

A DNA fragment containing 2,079 base pairs from Bacillus circulans CGMCC 1416 was cloned using degenerate PCR and inverse PCR. An open reading frame containing 981 bp was identified that encoding 326 amino acids residues, including a putative signal peptide of 31 residues. The deduced amino acid sequence showed the highest identity (68.1%) with $endo-{\beta}-1,4-D-mannanase$ from Bacillus circulans strain K-1 of the glycoside hydrolase family 5 (GH5). The sequence encoding the mature protein was cloned into the pET-22b(+) vector and expressed in Escherichia coli as a recombinant fusion protein containing an N-terminal hexahistidine sequence. The fusion protein was purified by $Ni^{2+}$ affinity chromatography and its hexahistidine tag cleaved to yield a 31-kDa ${\beta}$-mannanase having a specific activity of 481.55U/mg. The optimal activity of the purified protein, MANB48, was at $58^{\circ}C$ and pH 7.6. The hydrolysis product on substrate locust bean gum included a monosaccharide and mainly oligosaccharides. The recombinant MANB48 may be of potential use in the feed industry.

• Nutrition Research and Practice
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• v.6 no.1
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• pp.21-27
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• 2012

Pumpkins have considerable variation in nutrient contents depending on the cultivation environment, species, or part. In this study, the general chemical compositions and some bioactive components, such as tocopherols, carotenoids, and ${\beta}$-sitosterol, were analyzed in three major species of pumpkin ($Cucurbitaceae$ $pepo$, $C.$ $moschata$, and $C.$ $maxima$) grown in Korea and also in three parts (peel, flesh, and seed) of each pumpkin species. $C.$ $maxima$ had significantly more carbohydrate, protein, fat, and fiber than $C.$ pepo or $C.$ $moschata$ (P<0.05). The moisture content as well as the amino acid and arginine contents in all parts of the pumpkin was highest in $C.$ $pepo$. The major fatty acids in the seeds were palmitic, stearic, oleic, and linoleic acids. $C.$ $pepo$ and $C.$ $moschata$ seeds had significantly more ${\gamma}$-tocopherol than $C.$ $maxima$, whose seeds had the highest ${\beta}$-carotene content. $C.$ $pepo$ seeds had significantly more ${\beta}$-sitosterol than the others. Nutrient compositions differed considerably among the pumpkin species and parts. These results will be useful in updating the nutrient compositions of pumpkin in the Korean food composition database. Additional analyses of various pumpkins grown in different years and in different areas of Korea are needed.

• KSBB Journal
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• v.10 no.2
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• pp.149-158
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• 1995

The capacity of inclusion complex formation between ${\alpha}$-, ${\beta}$-, ${\gamma}$-cyclodextrins(CDs) and various compounds, such as pH indicators, biloslalns, glycoside, amino acid, and fatty acids, was compared. Fatty acid was identified as the most suitable ligand for fractionation of CDs in terms of capacity and selectivity. The effects of complex formation conditions, such as, mixing ratio of CD and fatty acid, pH, ionic strength, and temperature, on the capacity of fatty acrid-CD complex was also investigated. The carbon number of fatty acids was identified as the most significant factor determining the capacity and selectivity of inclusion complex formation of CDs. Capric acid(C10) and palmitic acid(C16) showed high specificity for ${\alpha}$- and ${\beta}$-CDs, respectively. Under the optimal conditions, the molar ratio of complex formed was found to be 1.0:2.6 for ${\alpha}$-CD/capric acid and 1.0:1.9 for ${\beta}$-CD/palmitic acid. X-ray diffraction and infrared spectrum of the formed inclusion complex were analyzed. The changes of enthalpy($\Delta$H) of the inclusion complex formation reaction was evaluated by differential scanning calorimetry, showed that the reaction was endothermic.

• Food Science of Animal Resources
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• v.28 no.4
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• pp.463-469
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• 2008

The objective of the present study was to compare the chemical, rheological and sensory properties of regular process cheese (control) and cholesterol-reduced block-type process cheese. The cholesterol-reduced process cheese was made by accelerated ripened cholesterol-reduced Cheddar cheese and cholesterol-reduced butter treated by crosslinked $\beta$-cyclodextrin ($\beta$-CD). The composition of the block-type process cheese was similar to the control cheese. Approximately 91.0% of cholesterol removal was observed when treated by crosslinked $\beta$-CD. The production of total free amino acids was significantly higher in cholesterol-reduced process cheese in all storage periods compared with those in the control. In sensory analysis, a significantly higher score of acidic, salty and bitterness. and lower score of elasticity were found. However, no difference was found in free fatty acids, rheological properties, and TBA test between the control and cholesterol-reduced process cheese. Therefore, the present study indicated that even though some of the significant difference was observed in sensory properties in the cholesterol-reduced block-type process cheese, most of chemical and rheological properties were comparable to the control process cheese.

• Korean Journal of Microbiology
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• v.45 no.2
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• pp.215-221
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• 2009

Site-specific incorporation of unnatural amino acids into proteins in vivo can be achieved by co-expression of an orthogonal pair of suppressor tRNA and engineered aminoacyl-tRNA synthetase (ARS) that specifically ligates an unnatural amino acid to the suppressor tRNA. As a step to establish this technique, here we generated an Escherichia coli reporter strain DH10B(Tn:lacZam) by integrating amber mutated lacZ gene into the chromosome of E. coli DH10B strain. In vivo expression of E. coli amber suppressor $tRNA^{Gln}$ produced blue colonies in culture plates containing X-Gal as well as dramatically increased $\beta$-galactosidase activity. In addition, expression of an orthogonal pair of Saccharomyces cerevisiae suppressor $tRNA^{Tyr}$ and tyrosyl-tRNA synthetase also produced blue colonies as well as moderate increase of $\beta$-galactosidase activity. These data demonstrate that our reporter strain will provide an efficient method to assess amber suppression in both qualitative and quantitative manners.

• Journal of Life Science
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• v.22 no.11
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• pp.1545-1551
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• 2012

Authors report the glycoside hydrolase (GH) family 16 ${\beta}$-agarase from the strain of Agarivorans sp. JA-1, which authors previously stated as recombinant expression and characterization of GH-50 and GH-118 ${\beta}$-agarase. It comprised an open reading frame of 1,362 base pairs, which encodes a protein of 49,830 daltons consisting of 453 amino acid residues. Valuation of the total sequence showed that the enzyme has 98% nucleotide and 99% amino acid sequence similarities to those of GH-16 ${\beta}$-agarase from Pseudoalteromonas sp. CY24. The gene corresponding to a mature protein of 429 amino acids was recombinantly expressed in Escherichia coli, and the enzyme was purified to homogeneity by affinity chromatography. It showed maximal activity at $40^{\circ}C$ and pH 5.0, representing 67.6 units/mg. Thin layer chromatography revealed that mainly neoagarohexaose and neoagarotetraose were produced from agarose. The enzyme would be valuable for the industrial production of functional neoagarooligosaccharides.