• Journal of Microbiology and Biotechnology
• /
• v.13 no.6
• /
• pp.912-918
• /
• 2003

Mitotic centromere-associated kinesin (MCAK), which is a novel kinesin with a central motor domain, is believed to playa role in mitotic segregation of chromosome during the M phase of the cell cycle. In the present study, it is shown that a rabbit polyclonal antibody has been produced using the N-terminal region (187 aa) of human MCAK expressed in E. coli as the antigen. To express the N-terminal region in E. coli, the MCAK cDNA fragment encoding N-terminal 187 aa was obtained by PCR and was then inserted into the pET 3d expression vector. Molecular mass of the N-terminal region overexpressed in the presence of IPTG was 23.2 kDa on SDS-PAGE, and the protein was insoluble and mainly localized in the inclusion body that could be easily purified from the other cellular proteins. The N-terminal region was purified by electro-elution from the gel after the inclusion body was resolved on the SDS-PAGE. The antiserum obtained after tertiary immunization with the purified protein specifically recognized HsMCAK when subjected to Western blot analysis, and showed a fluctuation of the protein level during the cell cycle of human Jurkat T cells. Synchronization of the cell-cycle progression required for recovery of cells at a specific stage of the cell cycle was performed by either hydroxyurea or nocadazole, and subsequent release from each blocking at 2, 4, and 7 h. Northern and Western analyses revealed that both mRNA and protein of HsMCAK reached a maximum level in the S phase and declined to a basal level in the G1 phase. These results indicate that a polyclonal antibody raised against the N-terminal region (187 aa) of HsMCAK, overexpressed in E. coli, specifically detects HsMCAK (81 kDa), and it can analyze the differential expression of HsMCAK protein during the cell cycle.

• Applied Biological Chemistry
• /
• v.46 no.4
• /
• pp.285-290
• /
• 2003

This study was conducted to select the bacteriocin-producing microoreanism cultivated in the rice bran culture and to characterize the produced bacteriocin for the further purpose of economical and anti-bacterial rice bran protein film. Pseudomonas putida 21025 was cultivated from rice bran and identified as a producer of a bacteriocin which showed bactericidal activity against Pseudomonas aeruginosa 9027. Bacteriocin produced by Pseudomonas putida 21025 showed a broad spectrum of activity against spoilage and soil bacteria. The activity of the bacteriocin produced by Pseudomonas putida 21025 decreased after 1 hr of staying at the temperature of $50^{\circ}C$, and with the presence of some organic solvents, except hexane and ethanol. However, the bacteriocin activity was stable throughout the pH ranges of 6-9 for 2 hrs, at the temperature lower than $50^{\circ}C$, and with the presence of ethanol for 3 hrs. The bacteriocin was partially purified by 50% ammonium sulfate precipitation followed by subsequent dialysis. Direct detection of the partially purified bacteriocin on SDS-PAGE suggested that it had an apparent molecular mass of about 21.6 kDa.

• Asian-Australasian Journal of Animal Sciences
• /
• v.31 no.2
• /
• pp.167-179
• /
• 2018

Objective: The aim of this study was to create a set of microsatellite markers with high polymorphism for the genetic monitoring and genetic structure analysis of local goose populations. Methods: Novel microsatellite markers were isolated from the genomic DNA of white Roman geese using short tandem repeated probes. The DNA segments, including short tandem repeats, were tested for their variability among four populations of geese from the Changhua Animal Propagation Station (CAPS). The selected microsatellite markers could then be used to monitor genetic variability and study the genetic structures of geese from local geese farms. Results: 14 novel microsatellite loci were isolated. In addition to seven known loci, two multiplex sets were constructed for the detection of genetic variations in geese populations. The average of allele number, the effective number of alleles, the observed heterozygosity, the expected heterozygosity, and the polymorphism information content were 11.09, 5.145, 0.499, 0.745, and 0.705, respectively. The results of analysis of molecular variance and principal component analysis indicated a contracting white Roman cluster and a spreading Chinese cluster. In white Roman populations, the CAPS populations were depleted to roughly two clusters when K was set equal to 6 in the Bayesian cluster analysis. The founders of private farm populations had a similar genetic structure. Among the Chinese geese populations, the CAPS populations and private populations represented different clads of the phylogenetic tree and individuals from the private populations had uneven genetic characteristics according to various analyses. Conclusion: Based on this study's analyses, we suggest that the CAPS should institute a proper breeding strategy for white Roman geese to avoid further clustering. In addition, for preservation and stable quality, the Chinese geese in the CAPS and the aforementioned proper breeding scheme should be introduced to geese breeders.

• Asian Pacific Journal of Cancer Prevention
• /
• v.15 no.3
• /
• pp.1363-1367
• /
• 2014

This research was conducted to compare differences in colon cancer lymphatic vessel invasion (LVI) with D2-40 antibody labeling and regular HE staining, blood vessel invasion (BVI) with CD34 antibody labeling and HE staining and to assess the possibility of using D2-40-LVI/CD34-BVI in combination for predicting stage II colon cancer prognosis and guiding adjuvant chemotherapy.Anti-D2-40 and anti-CD34 antibodies were applied to tissue samples of 220 cases of stage II colon cancer to label lymphatic vessels and small blood vessels, respectively. LVI and BVI were assessed and multivariate COX regression analysis was performed for associations with colon cancer prognosis. Regular HE staining proved unable to differentiate lymphatic vessels from blood vessels, while D2-40 selectively labeled lymphatic endothelial cell cytosol and CD34 was widely expressed in large and small blood vessels of tumors as well as normal tissues. Compared to regular HE staining, D2-40-labeling for LVI and CD34-labeling for BVI significantly increased positive rate (22.3% vs 10.0% for LVI, and 19.1% vs 9.1% for BVI). Multivariate analysis indicated that TNM stage, pathology tissue type, post-surgery adjuvant chemotherapy, D2-40-LVI, and CD34-BVI were independent factors affecting whole group colon cancer prognosis, while HE staining-BVI, HE staining-LVI were not significantly related. When CD34-BVI/D2-40-LVI were used in combination for detection, the risk of death for patients with two or one positive results was 5.003 times that in the LVI(-)&BVI(-) group (95% CI 2.365 - 9.679). D2-40 antibody LVI labeling and CD34 antibody BVI labeling have higher specificity and accuracy than regular HE staining and can be used as molecular biological indicators for prognosis prediction and guidance of adjuvant chemotherapy for stage II colon cancer.

• Journal of the Society of Cosmetic Scientists of Korea
• /
• v.38 no.1
• /
• pp.43-50
• /
• 2012

The safety of cosmetics or cosmetic ingredients on human skin is generally evaluated by visual assessment but some early subtle skin changes may not be noticed by the naked eyes. Thus, the present study was conducted to detect skin reactions induced by mildly irritating cosmetic ingredients by using a laser Doppler perfusion imager (LDPI) method that measures blood flow, a $Vapometer^{(R)}$ that measure strans epidermal water loss (TEWL), and a spectrophotometer that measures the skin color as the erythema values ($a^*$). Visual assessment showed that all tested oils and humectants except propylene glycol belong to the low skin irritation ranges (grades 0+ to 2.9+) while all tested surfactants and propylene glycol belong to the moderate-to strong-skin irritation ranges (grades 3+ to 5+). Among three instrumental methods, TEWL assessment appeared to be more sensitive than spectrophotometric or LDPI method and suitable for the detection of subtle skin response invisible to the naked eye (grades 0+ to 2.9+). Skin reactions of grade 3+ to 5+ could be detected by all three instrumental methods. In conclusion, the current study suggested that the sub-clinical skin reactions due to mild irritants contained in cosmetics can be best assessed by TEWL measurements.

• Journal of the Society of Cosmetic Scientists of Korea
• /
• v.40 no.4
• /
• pp.359-364
• /
• 2014

In this study, we developed a HPLC method for the separation and analysis of methylparaben and p-anisic acid, which are commonly used as a synthetic preservative and natural preservative, respectively. Methylparaben and p-anisic acid have same molecular weight (152.15 g/mol), similar structure and same maximum absorption wavelength (250 nm), thus they showed same retention time (RT) value (13.3 min) in HPLC experiment. We observed that two substances are separated on C18 column after methylparaben was derivatized selectively through the acetylation reaction. Instead, RT of the acetylated methylparaben was moved to 23.9 min from 13.3 min. The average retention time was $23.9{\pm}0.1min$ and peak area values was $5042882{\pm}4778$. In addition it showed a high linearity in the calibration curve with a correlation coefficient (R2) of 0.9999658. Detection and quantitation limits were $1.47{\mu}g/mL$ and $4.44{\mu}g/mL$, respectively. In conclusion, the developed method can be useful for separation and analysis of preservatives with similar structure in cosmetic fields.

• Korean Journal of Microbiology
• /
• v.48 no.2
• /
• pp.116-124
• /
• 2012

Identification was performed in March 2008 for the 76 Enterococcus strains isolated from the Han-river, which is used as water supply for Seoul citizens. The antibiotic susceptibility, antibiotic resistant structural analysis, trans-conjugation, pulsed-field gel electrophoresis (PFGE) and multi-locus sequence typing (MLST) were also carried out for the isolated strains. Among the isolated strains, 25 strains were E. casseliflavus, 4 strains were E. faecalis and 1 strain was E. hirae. Investigation of antibiotic susceptibility indicated that 15 strains demonstrated tolerance against vancomycin, and that 11 strains of E. faecium and 4 strains of E. casseliflavus were VRE. The vanA gene detection of the VRE strains revealed that 6 E. faecium strains were vancomycin-resistant Enterococcus faecium (VREF) possessing vanA. Analyses of transposon Tn1546 structure containing vanA demonstrated that Km36 and Km37 belonged to Tn1 type, Km20 and Km38 was Tn2 type, and Km39 and Km40 was Tn3 type. PFGE disclosed that among the 6 VREF strains, Km36 and Km37 exhibited equivalent subtype, while the rest 4 strains showed subtypes different to each other. MLST for the 6 VREF strains disclosed that 3 strains were ST78, while the rest 3 strains were ST18, ST192 and ST230, respectively. All these clonal complexes were derived from CC17 which has been isolated from clinical sources. 4 strains belonged to CC78, while the rest 2 strains were CC18 and CC192, respectively.

• Journal of Life Science
• /
• v.20 no.8
• /
• pp.1281-1286
• /
• 2010

Gaucher disease (GD) is caused by glucocerebrosidase functional deficiency and the most prevalent lysosomal storage disorder (LSD), with an incidence of about 1 in 20,000 new births. Resveratrol, one kind of phytoalexin, is a produced naturally by several plants and has anti-tumor, anti-aging, anti-inflammatory and neuro-protective effects. In this paper we provide the cellular protective effect of resveratrol in both type I and type II Gaucher disease cells. Resveratrol treatment did not show any significant change in the p21 and p53 mRNA expression level, however expression level of the p21 protein, a cell cycle arrest factor, shows significant increment in both types of Gaucher disease cells. These cell cycle arrest patterns were confirmed by both MTT assay measurement and microscopy detection. In comparison, expression level of poly ADP ribose polymerase (PARP), an apoptosis indicator protein, was significantly decreased in both type I and II Gaucher disease cells after treatment with resveratrol. This result indicates that resveratrol relievescellular apoptotic stress fromtype I and II Gaucher disease cells. Therefore, we demonstrate that resveratrol inhibits cell proliferation via p21 activity and activates cellular repair systems for Gaucher disease cells. Our results provide at least one of the molecular mechanisms of Gaucher disease and may allow the verification of potential drug targets for therapeutic trials.

• Journal of Periodontal and Implant Science
• /
• v.46 no.5
• /
• pp.320-328
• /
• 2016

Purpose: Human saliva, as a vital part of the immune defense system, contains a number of distinct proteins and peptides. Recently human common salivary protein 1 (CSP1) has been identified as an abundant salivary protein and may play a role in promoting the binding of cariogenic bacteria to salivary pellicles. However, nothing else is known regarding the role of CSP1 in periodontology. The aim of this study was to quantify and compare CSP1 levels between healthy subjects and periodontal patients. Methods: This controlled clinical study was conducted in periodontally healthy individuals and patients with chronic periodontitis Chonbuk National University Hospital, with Institutional Review Board approval. Whole saliva samples were collected from 36 healthy subjects and 33 chronic periodontitis patients and analyzed. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immune blotting were conducted to ensure that anti-CSP1 monoclonal antibody (mAb) binds to CSP1 in human saliva. A sandwich enzyme-linked immunosorbent assay (ELISA) system was house-fabricated using mAb-hCSP1#14 and mAb-hCSP1#4 as a capture and a detector mAb, respectively. The CSP1 concentrations in saliva from 36 healthy subjects and 33 periodontal patients were quantified using the CSP1 sandwich ELISA system, and the results were analyzed using the Student's t-test. Results: Immunoblot analysis using mAb-hCSP1 as a probe confirmed that CSP1 in human saliva existed as a single band with a molecular weight of approximately 27-kDa. The quantification of CSP1 concentrations by CSP1 ELISA showed that the median values (25th to 75th percentiles) of periodontal patients and healthy subjects were 9,474 ng/mL (range, 8,434.10,139 ng/mL) and 8,598 ng/mL (range, 7,421.9,877 ng/mL), respectively. The Student's t-test indicated the presence of a statistically significant difference between the 2 groups (P=0.024). Conclusions: The presence of a significant difference in CSP1 levels between healthy subjects and periodontal patients suggests that CSP1 may be a potential biomarker for the detection or screening of periodontitis patients.

• Pediatric Infection and Vaccine
• /
• v.27 no.2
• /
• pp.90-101
• /
• 2020

Purpose: The multiplex reverse transcriptase-polymerase chain reaction (RT-PCR) test developed recently can help detect enteric pathogens of acute gastroenteritis (AGE). This study aimed to investigate the epidemiology of pathogens in children with AGE using the multiplex RT-PCR. Methods: From May 2015 to June 2018, multiplex RT-PCR tests were performed to identify pathogens in the feces of pediatric patients diagnosed with AGE at a secondary hospital in Seoul, Korea. Results: Of the 1,366 stool samples examined for viral pathogens, 483 (35.3%) tested positive for ≥1 pathogen. Group A rotavirus (RV) was detected in 106 cases (7.8%). The positivity rate increased annually from 3.0% (8/263) to 16.7% (48/288) and surged in 2018 (P<0.001). Norovirus (NoV) GII was the most common viral pathogen (263/1,366, 19.3%), and the positivity rate did not increase during the 3 years. Of the 304 stool samples tested for bacterial pathogens, Campylobacter spp. was the most common bacterial pathogen (32/304, 10.5%), followed by Clostridium difficile (22/304, 7.2%) and Salmonella spp. (17/304, 5.6%). The positivity rate of these bacterial pathogens did not change significantly during the study period. Conclusions: NoV GII is the main pathogen in childhood AGE since the introduction of RV vaccine, yet the number of rotavirus-infected patients increased during our study, especially in 2018. Therefore, further research is needed including the possibility of emergence of novel RV strains. Campylobacter spp. is the predominant cause of bacterial AGE in children. For proper treatment, the clinical characteristics of the bacteria should be taken into consideration, and continuous monitoring is necessary.