• Restorative Dentistry and Endodontics
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• v.20 no.2
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• pp.645-654
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• 1995

Porphyromonas endodontalis is a black-pigmented anaerobic Gram negative rod which is associated with endodontal infections. It has been isolated from infected dental root canals and submucous abscesses of endodontal origin. DNA probe is an available alternative, offering the direct detection of a specific microorganism. Nucleic-acid probes can be off different types: whole different: whole-genomic, cloned or oligonucleotide probes. Wholegenomic probes are the most sensitive because the entire genome is used for possible hybridization sites. However, as genetically similar species of bacteria are likely to be present in specimences, cross-reactions need to be considered. Cloned probes are isolated sequences of DNA that do not show cross-reactivity and are produced in quantity by cloning in a plasmid vector. Cloned probes can approach the sensitivity found with whole-genomic probes while avoiding known cross-reacting species. Porphyromonas endodontalis ATCC 35406 (serotype $O_1K_1$) was selected in this experiment to develop specific cloned DNA probes. EcoR I-digested genomic DNA fragments of P. endodontalis ATCC 35406 were cloned into pUC18 plasmid vector. From the E. coli transformed with the recombinant plasmid 4 clones were selected to be tested as specific DNA probes. Restriction-digested whole-genomic DNAs prepared from P. gingivalis 38(serotype a), W50(serotype b), A7A1-28(serotype C), P. intermedia 9336(serotype b), G8-9K-3(serotype C), P. endodontalis ATCC 35406(serotype $O_1K_1$), A. a Y4(serotype b), 75(serotype a), 67(serotype c), were each seperated on agarose gel electrophoresis, blotted on nylon membranes, and were hybridized with digoxigenin-dUTP labeled probe. The results were as follows: 1. Three clones of 1.6kb(probe e), 1.6kb(probe f), and 0.9kb(probe h) in size, were obtained. These clones were identified to be a part of the genomic DNA of P. endodontalis ATCC 35406 judging from their specific hybridization to the genomic DNA fragments of their own size on Southern blot. 2. The clones of 4.9kb(probe i) was identified to be a part of the genomic DNA of P. endodontalis ATCC 35406. but not to specific for itself. It was hybridized to P. gingivalis A7A1-28, P. intermedia G89K-3.

• The Korean Journal of Physiology
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• v.17 no.2
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• pp.177-182
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• 1983

In order to clarify the effect of the unilateral pneumonectomy on the compensatory growth of the residual lung, liver and kidney, and serum electrophoresis pattern, right lung pneumonectomy was performed on rabbits under general anesthesia with pentobarbital sodium. On the fifth day after the surgery, the weight of the residual lung, liver and bilateral kidneys was measured and organ weight-body weight ratio was calculated. And in an attempt to know whether the cells in the liver and the kidney were proliferated by unilateral pneumonectomy, DNA content was determined. The quantity(g/100 ml) of serum protein was determined also and serum electrophoresis was performed on cellulose acetate membrane. The results obtained are summarized as following. The weight of the residual lung and lung weight-body weight ratio was significantly increased respectively. The weight of the liver and organ weight-body weight ratio were not changed but the DNA content of the liver and kidney tissue increased significantly, illustrating that unilateral pneumonectomy caused cellular hyperplasia in the liver and in the kidney as well as in the residual lung. The quantity(g/100 ml) of serum protein was significantly increased and in the analysis of the electrophoregram, there was significant difference between the normal and pneumonectomy group. Taken together, these results indicate that unilateral pneumonectomy caused the compensatory hyperplasia of the liver, the kidney and the residual lung as well as the change of electro-phoretic pattern. And it also suggests that a humoral factor, which proliferates the cells in the residual lung, the liver and the kidney, existed in the pneumonectomized rabbits.

• Korean Journal of Clinical Laboratory Science
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• v.48 no.4
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• pp.371-377
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• 2016

Decalcification is routinely performed to obtain a pathological diagnosis using bone marrow biopsy. During the decalcification process using a conventional acidic solution, such as HCl, the antigenicity of tissue is damaged. Especially DNA and RNA in the bone marrow are impaired. Hence, there is the need for a standardized decalcification protocol that preserves the antigenicity of tissue. To this end, we compared the effects of two commonly used decalcifiers: Commercial decalcifier (Calcl-Clear Rapid, HCl) and the EDTA (12.5%, pH 7.0). Bone marrow biopsies sampled from 71 patients were decalcified in accordance with the protocols of respective groups-HCI versus EDTA. The differences of decalcification protocols were analyzed with respect to Hematoxylin & Eosin staining, Gomori'sreticulum staining, and immunohistochemical staining and molecular analysis. Immunohistochemical staining used Ki-67, CD20 and CD138 as primary antibodies and molecular analysis was conducted through the DNA concentration analysis, in situ hybridization (ISH) and immunoglobulin heavy chain (IGH) gene rearrangement. On the routine histopathology analysis, there was no difference between HCl and EDTA. Moreover, in case of immunohistochemical staining, the cytoplasmic membrane or cytoplasmic CD markers was well preserved. However, nuclear proteins, such as Ki-67, were stained with low quality. Conversely, according to the molecular analysis, the EDTA protocol preserved the DNA and RNA compared with the HCI. The differences of DNA quantity and quality were statistically significant between protocols of HCl and EDTA. We used 38 cases in HCl and 12 cases in EDTA. Consequently, the EDTA protocol maintains the antigenicity of the protein on tissue and is acceptable for examination with molecular base analysis. Decalcification of bone marrow biopsy by EDTA is highly recommended for the examination of immunohistochemical staining and molecular analysis.

• Korean Journal of Poultry Science
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• v.40 no.1
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• pp.31-40
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• 2013

This study was performed to investigate the investigated effects of dietary supplementation of vitamin C and E on the growth performance and stress response in broiler chickens. Stress response was analyzed by the quantity of telomeric DNA, the rate of DNA damage and the expression levels of heat shock proteins (HSPs) and hydroxyl-3-methyl-glutaryl coenzyme A reductase (HMGCR) genes on tissues and blood. The telomere length and telomere shortening rates were analyzed by quantitative fluorescence in situ hybridization on the nuclei of lymphocytes and tissues. The DNA damage rate of lymphocytes was quantified by the comet assay. The expression levels of HSP70, HSP90s and HMGCR genes were measured by quantitative real-time polymerase chain reaction in lymphocytes. In results, there was no significant difference among treatments in body weight, weight gain, feed intake and mortality. The telomere shortening rate of the lymphocytes was significantly lower in the vitamin E supplemented group than the control group. The DNA damage was also decreased supplemented with vitamin C and E, as compared to the control group. The vitamin E supplemented group had a significant positive effect on the expressions of HMGCR, HSP90-${\alpha}$ and HSP90-${\beta}$ in lymphocytes, but had no significance on HSP70, as compared to the control group. We concluded that the dietary supplementation of vitamin E (100 mg/kg feed) had reduced the individual physiological stress response without stunt growth in broiler chickens.

• Korean Journal of Poultry Science
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• v.36 no.4
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• pp.293-300
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• 2009

The rate of fetus with abnormal chromosomes increase with maternal age. Nondisjunction of aging oocyte chromosome is a major reason for the increased rate of abnormalities. Telomeres are the ends of eukaryotic chromosome, which are essential for chromosome stability and are related in cell senescence. This study was carried out to analyze the chromosome aberration rate and amount of telomeric DNA in chick embryo along with maternal age. Fertilized eggs and blood were sampled from White Leghorn layers starting at 20 weeks through to 70 weeks age at 10 weeks interval. Chromosome aberration rate was analyzed by karyotyping. The amounts of telomeric DNA in embryonic cells and lymphocytes were quantified by Quantitative Fluorescence in situ Hybridization method. The chromosome aberration rate in chick embryos significantly differed with maternal age. The chromosome aberration rate increased at early laying period and beyond 70 weeks of maternal age. Therefore, chromosome aberration rate was affected by maternal age due to ovulated oocytes state. However, the amount of telomeric DNA on embryonic cells did not differ significantly with maternal age. Thus, maternal age does not affects telomere quantity in their embryos due to cellular reprograming at early embryonic stage after fertilization.

• Environmental Mutagens and Carcinogens
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• v.24 no.2
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• pp.79-84
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• 2004

Three synthetic chemicals, benzoyl chloride, 2-propyn-l-ol, and 2-phenoxy ethanol were selected for genotoxicity testing, based on production quantity and available genotoxic data. In our previous report, benzoyl chloride induced chromosomal aberrations in Chinese hamster lung (CHL) fibroblast in vitro with and without metabolic activation, while 2-propyn-l-ol and 2-phenoxy ethanol induced only with metabolic activation. To compare the genotoxicity of chromosome aberration assay, the single cell gel electrophoresis (comet) assay subjected using CHL cells. As a result, statistically significant differences of tail moment values of benzoyl chloride, 2-propyn-1-ol, and 2-phenoxy ethanol were observed compared with control values on almost all concentrations with S9 or without S9 metabolic activation system. This results suggest that genotoxic results of the comet assay and the chromosome aberration assay show correlationship of genotoxicity in the CHL fibroblast. In summary, the positive result of chromosome aberration of benzoyl chloride, 2-propyn-l-ol, and 2-phenoxy ethanol was also induced DNA damages in comet assay with same cell line. Consequently, comet assay will be useful and more accurate tool to detect and to confirm the genotoxicity especially DNA damages in CHL fibroblast.

• YAKHAK HOEJI
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• v.45 no.1
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• pp.71-77
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• 2001

We recently developed a high efficiency expression vectors pCK, which drives a high level of gene expression in the skeletal muscles of mice. In this study, we investigated the pharmacokinetics and biodistribution of pCK-VEGF expressing human VEGF165 after intravenous or intramuscular administration. The quantity of pCK-VEGF in the tissues of mice was measured by the PCR method which has a detection limit of approximately 1 pg of the exogenously added plasmid. In the case of intravenous administration, the half life of the pCK-VEGF plasmid in the bloodstream was 1.68 min. After intra-muscular administration, the half life of pCK-VEGF plasmid in the bloodstream was 6.78 min. At 90 min post-administration, 30% of the injected pCK-VEGF was found at the site of injection, where it persisted for up to 8 hours. Less than 1.6% of the injected pCK-VEGF plasmid DNA was detected in highly vascularized tissues such as the lung, kidney; and liver at 90 min post-administration, but the plasmid was undetectable at later time points. These results suggested that intramuscularly administrated pCK-VEGF persisted for longer periods of time in muscles than in other tissues and that direct intra-muscular injection of pCK-VEGF might be useful for local therapeutic angiogenesis.

• Journal of the Korean Electrochemical Society
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• v.5 no.4
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• pp.202-208
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• 2002

We have been investigating electropolymerization of pyrrole in aqueous electrolyte solutions in acidic as well as in neutral conditions by in situ electrochemical quartz crystal oscillator method, where resonant frequency and resonant resistance can be monitored simultaneously with current-voltage measurements during electropolymerization of pyrrole. The properties of thin PPy films prepared on electrode surfaces depended strongly on the experimental variables of electrode potentials applied, solution pH, kinds and quantity of supporting electrolytes, added chemicals, and the mode of electrochemical method employed. We are applying our experience gained on electropolymerization of pyrrole to immobilizing biomolecules onto electrode surfaces to develop a biosensor system. In this work, we wish to present the results on electrochemical monitoring on electropolymerization of pyrrole in the presence of DNA and albumin in different electrochemical conditions. Additionally we will summarize our investigations on the miniaturization of biomolecules/PPy composites by means of scanning tunneling microscopy.

• Journal of Korean society of Dental Hygiene
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• v.24 no.2
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• pp.131-139
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• 2024

Objectives: Oral bacterial samples included subgingival, supragingival, and saliva plaques. As the diversity and number of microorganisms deffer depending on the area of the oral cavity and the method used, an appropriate and reliable collection method is important. The present study investigated oral bacterial sampling methods. Methods: Supragingival dental plaque was collected from the buccal and lingual tooth surfaces of study participants using sterilized cotton swabs. Plaques were collected from the subgingival area using a sterilized curette. Bacterial genomic DNA was extracted using MagNA Pure 96 DNA and Viral NA low-volume kits. Real-time polymerase chain reaction (PCR) was performed using the PowerCheck^TM Periodontitis Pathogens Multiplex Real-time PCR kit. Results: Aggregatibacter actinomycetemcomitans, Prevotella intermedia, and Fusobacterium nucleatum of the orange complex were not observed in the subgingival biofilms of all study participants. For Porphyromonas. gingivalis, a significant correlation was observed between supragingival, subgingival, and total tooth surface biofilms. Compared to the supragingival and subgingival biofilmss, total tooth surface biofilm exhibited the highest bacterial count when the inswabbing method was used. Conclusions: Based on these findings, the supragingival swab method is recommended for oral bacterial research.

• Reproductive and Developmental Biology
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• v.29 no.1
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• pp.1-7
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• 2005

Telomeres consisting of (TTAGGG)n tandem repeat DNA sequences and associated proteins are essential for chromosome stability and related with cell senescence, apoptosis and cancer. The telomerase is a ribonucleoprotein which act as a template for the synthesis of telomeric DNA. This study was carried out to identify the distribution of telomeres on mouse chromosomes and also to analyze the amount of telomeres and telomerase activity of mouse embryos at early embryonic stages. Germ cells and early embryos from 1 cell to blastocyst stage were analyzed. The amount of telomeres was analyzed by quantitative fluorescence in situ hybridization technique(Q-FISH) using a human telomeric DNA probe, and telomerase activity was measured by telomeric repeat amplification protocol assay(TRAP). In results, the telomeres on mouse chromosomes were distributed at the ends of all autosomes and sex chromosomes. Although the quantity of telomeres varied among chromosomes, most of chromosomes had higher amount in q-arm telomeres than in p-arm telomeres. The results of Q-FISH indicated that the relative amount of telomeres of mouse embryos in each embryonic stage was approximately the same except the higher amount in blastocysts. Using TRAP assay on mouse embryos, telomerase activity was detected in all preimplantation stages from mature oocytes to blastocysts. Especially the telomerase activity was significantly increased at the morula and blastocyst stage. In conclusion, there may be a close association between the amount of telomeres and telomerase activity in early embryonic stages, and analysis of telomere quantity and telomerase activity on early development will be helpful for the investigation of embryogenesis and embryonic cell differentiation in mice.