• Title/Summary/Keyword: DNA assay

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Evaluation on the Applicability of Comet Assay for the Identification of Gamma-irradiated Frozen Beef Cut, Ground Beef and Tteokgalbi (감마선처리 냉동 우육, 분쇄우육 및 떡갈비의 판별을 위한 Comet assay의 적용성 평가)

  • Jong-Heum Park;Jae-Kyung Kim;Beom-Seok Song
    • Journal of Radiation Industry
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    • v.17 no.3
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    • pp.257-264
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    • 2023
  • This study was conducted to evaluate the applicability of the Comet assay, which is widely used for the identification of irradiated meats, to detect irradiated beef cut, ground beef, and Tteokgalbi during freezing storage. Gamma-irradiation significantly increased the DNA damage in frozen beef cut and ground beef samples. Among those, DNA nuclei of samples irradiated with absorbed doses of 1kGy or more showed typical comet-shaped damage, convincing that the samples were irradiated. Meanwhile, DNA nuclei in non-irradiated beef cut and ground beef samples were also damaged according to storage time. In particular, since the damage of DNA nuclei in the non-irradiated samples frozen for three months was similar to that of samples irradiated with a dose of 0.5 kGy, it was considered difficult to detect whether these samples were irradiated by Comet assay analysis. Likewise, gamma-irradiation of Tteokgalbi increased DNA damage. However, significant damage to DNA nuclei was observed even in the non-irradiated samples. Therefore, the application of the analysis method to determine whether the Tteokgalbi sample was irradiated was not appropriate. In conclusion, these results suggest that Comet assay could be limitedly applied only to fresh meat with a short storage period and minimal processing.

In vivo Comet Assay on Flounder and Clam Exposed to BaP and TBT (BaP 및 TBT에 노출된 넙치와 개조개의 in vivo Comet assay)

  • Kim, So-Jung;Chung, Young-Jae;Lee, Taek-Kyun
    • Ocean and Polar Research
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    • v.33 no.2
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    • pp.127-133
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    • 2011
  • The comet assay, also called single-cell electrophoresis (SCGE) assay, is a potential sensitive monitoring tool for DNA damage in cells. The primary objective of this study was to use comet assay to ascertain if the blood cells of flounder (Pleuronichthys olivaceus) and muscle cells of clam (Saxidomus purpurata) are suitable for genotoxicity screening. This was achieved by initially exposing blood and muscle cells under in vitro conditions to the reference genotoxin hydrogen peroxide ($H_2O_2$); strong correlation between $H_2O_2$ concentration and comet values were found. Subsequently, the identification of DNA damage in isolated cells from flounder and clam was performed under in vivo exposure to benzo(a)pyrene (BaP) and tributyltin (TBT). Flounder and clam were exposed to different concentrations (1, 10, 50, 100 ${\mu}g/L$) of BaP or TBT for 4 days. Regardless of treated chemicals, blood cells of flounder were more prone to DNA breakage compared to muscle cells of clam. In conclusion, in vivo genotoxicity of BaP and TBT can be biomonitored using the comet assay. This study suggests that flounder and clam do show potential as mediums for monitoring genotoxic damage by comet assay.

Rapid Analysis of Genetic Relationship of Phytoplasma Isolates by a DNA Heteroduplex Mobility Assay (DNA Heteroduplex Mobility Assay법을 이용한 파이토플라스마 병원체의 유연관계 분석)

  • ;Chuji Hiruki
    • Korean Journal Plant Pathology
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    • v.14 no.5
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    • pp.382-385
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    • 1998
  • Molecular identification and genetic relationships between a phytoplasma associated with chestnut little leaf (CLL) and phytoplasma isolates of other trees in Korea were amplified by polymerase chain reaction (PCR). These 16S rDNA sequences amplified from the various phytoplasmas were used in DNA heteroduplex mobility assays (HMA). In DNA HMA combined with PCR, the mobility shift was observed for a heteroduoplex formed in combined with CLL and jujube witches broom, but not for those formed in combined with CLL and each of sumac witches broom, paulownia witches broom, and mulberry dwarf. HMA combined with PCR has been shown to be a very useful method for detection and differentiation of phytoplasmas.

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Application of the SCGE Assay for Detecting Induced DNA Damage in Plant Leaves

  • Kim, Jin Kyu;Song, Hi Sup;Kim, Do Young;Gichner, Tomas
    • Proceedings of the Korea Society of Environmental Biology Conference
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    • 2003.11a
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    • pp.68-73
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    • 2003
  • The possibility of using the alkaline protocol of the single cell gel electrophoresis (SCGE) assay as a method for detecting induced DNA damage has been studied for six major plants. The EMS was applied as a model genotoxic agent on young excised leaves of the tested crops for 18 h at 26$^{\circ}C$ in the dark. With increasing concentrations of 0 to 10 mM EMS, the DNA damage, expressed by the averaged median tail moment values, significantly increased in nuclei of all plants studied. As the results, no correlation between the diameter of nuclei and sensitivity to EMS treatment was observed. The data obtained demonstrate the feasibility of using the SCGE assay for detecting induced DNA damage in plants.

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DNA Damage of Lymphocytes in Volunteers after 4 hours Use of Mobile Phone (일부 자원자들의 이동전화 4시간 연속 사용 후 림프구 DNA 손상 평가)

  • Ji, Seon-Mi;Oh, Eun-Ha;Sul, Dong-Geun;Choi, Jae-Wook;Park, Hee-Chan;Lee, Eun-Il
    • Journal of Preventive Medicine and Public Health
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    • v.37 no.4
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    • pp.373-380
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    • 2004
  • Objectives : There has been gradually increasing concern about the adverse health effects of electromagnetic radiation originating from cell phones which are widely used in modern life. Cell phone radiation may affect human health by increasing free radicals of human blood cells. This study has been designed to identify DNA damage of blood cells by electromagnetic radiation caused by cell phone use. Methods : This study investigated the health effect of acute exposure to commercially available cell phones on certain parameters such as an indicator of DNA damage for 14 healthy adult volunteers. Each volunteer during the experiment talked over the cell phone with the keypad facing the right side of the face for 4 hours. The single cell gel electrophoresis assay (Comet assay), which is very sensitive in detecting the presence of DNA strand-breaks and alkali-labile damage in individual cells, was used to assess peripheral blood cells (T-cells, B-cells, granulocytes) from volunteers before and after exposure to cell phone radiation. The parameters of Comet assay measured were Olive Tail Moment and Tail DNA %. Results : The Olive Tail Moment of B-cells and granulocytes and Tail DNA % of B-cells and granulocytes were increased by a statistically significant extent after 4-hour use of a cell phone compared with controls. Conclusion : It is concluded that cell phone radiation caused the DNA damage during the 4 hours of experimental condition. Nonetheless, this study suggested that cell phone use may increase DNA damage by electromagnetic radiation and other contributing factors.

Comparison of lymphocyte DNA damage levels and total antioxidant capacity in Korean and American diet

  • Lee, Min Young;Kim, Hyun A;Kang, Myung Hee
    • Nutrition Research and Practice
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    • v.11 no.1
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    • pp.33-42
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    • 2017
  • BACKGROUND/OBJECTIVE: This study aims to measure the in vitro antioxidant capacity of Korean diet (KD) with American diet (AD) as a control group and to examine the ex vivo DNA damage reduction effect on human lymphocytes. MATERIALS/METHODS: The KD applied in this study is the standard one-week meals for Koreans (2,000 kcal/day) suggested by 2010 Dietary Reference Intakes for Koreans. The AD, which is the control group, is a one-week menu (2,000 kcal/day) that consists of foods that Americans would commonly take in according to the National Health and Nutrition Examination Survey. The antioxidant capacity of each menu was measured by means of the total phenolic assay and 3 in vitro antioxidant activity assays (2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity, trolox equivalent antioxidant capacity (TEAC), Oxygen radical absorbance capacity ($ORAC_{ROO{\cdot}}$)), while the extent of ex vivo lymphocyte DNA damage was measured by means of the comet assay. RESULTS: When measured by means of TEAC assay, the in vitro antioxidant capacity of the KD of the day was higher than that of the AD (P < 0.05) while there was no significant difference in total phenolic contents and DPPH and ORAC assays. The ex vivo lymphocyte DNA damage protective effect of the KD was significantly higher than that of the AD (P < 0.01). As for the one-week menu combining the menus for 7 days, the total phenolic assay (P < 0.05) and in vitro antioxidant capacity (P < 0.001, DPPH; P < 0.01, TEAC) of the KD menu were significantly higher than those of the AD menu. Likewise, the ex vivo DNA damage reduction rate of the Korean seven-day menu was significantly higher than that of the American menu (P < 0.01). CONCLUSION: This study demonstrates that the high antioxidant capacity and DNA damage protective effect of KD, which consists generally of various plant foods, are higher than those of typical AD.

Identification Characteristics of Irradiated Dried Red Pepper during Storage by Analysis of Thermoluminescence, DNA Comet, and DEFT/APC (Thermoluminescence, DNA Comet 및 DEFT/APC 분석에 의한 방사선처리 건고추의 저장 중 검지 특성)

  • Kim, Byeong-Keun;Kwon, Joong-Ho
    • Korean Journal of Food Science and Technology
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    • v.36 no.6
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    • pp.851-856
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    • 2004
  • Minerals separated from irradiated dried red pepper (whole) at 2.5 kGy or higher showed typical thermoluminescence (TL) glow curves ($TL_1$) at around $150^{\circ}C$, which increased with irradiation dose. The TL ratio ($TL_1/TL_2$) through re-irradiation step at 1 kGy enhanced reliability of TL identification results. DNA comet assay indicated that the intact cell was observed in non-irradiated pepper (seed), while some long tails were found in irradiated ones, showing relationship between irradiation dose and tail length. Log DEPT/APC values increased in proportion to irradiation doses in powdered and whole peppers. Based on overall results, irradiated dried red peppers could be screened using DNA comet assay or log DEFT/APC, and moreover the identification results were verified by TL analysis.

Screening of In vitro DNA-binding Activity from the Crude Drugs (생약으로부터 In vitro DNA 결합활성의 검색)

  • Kim, Youn-Seol;Jeong, Sei-Joon;Shin, Hwa-Woo;Kim, Youn-Chul
    • YAKHAK HOEJI
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    • v.42 no.2
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    • pp.144-148
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    • 1998
  • One hundred and seventeen crude drugs were screened for DNA-binding activity in vitro. The DNA-methyl green assay is a useful biochemical screen for the detection or development of biologically active compounds which bind to DNA. This assay is based upon the fact that free methyl green undergoes rapid spontaneous molecular rearrangement to its colorless carbinol base so that the liberation of the dye from DNA by displacement can be followed spectrophotometrically as a decrease in absorbance at 65Onm. Seven methanolic extracts of crude drugs including Cinnamomi Cortex spissus, Coicis Semen, Coptidis Rhizoma, Perillae Semen, Plantaginis Semen, Polygalae Radix and Zanthoxyli Fructus showed less than 2,000${\mu}$g/ml as their $IC_{50}$ values. The active principles of Plantaginis Semen and Zanthoxyli Fructus were transferred into organic solvents, which showed the $IC_{50}$ values with 588 and 574${\mu}$g/ml, respectively. These fractions have been selected for isolation of biologically active constituents.

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Rapid detection and quantification of porcine circovirus type 2 (PCV 2) DNA in Real-time PCR (Real-time PCR을 이용한 돼지써코바이러스 감염증 진단법 연구)

  • Kim, Eun-Gyeong;Hwang, Bo-Won;Lee, Jong-Min;Son, Byeong-Guk;Park, Ho-Jung;Kim, Tho-Kyoung
    • Korean Journal of Veterinary Service
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    • v.32 no.4
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    • pp.299-306
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    • 2009
  • Assay for the detection and quantification of porcine circovirus type 2 (PCV 2) with the real-time PCR were developed. TaqMan probe real-time using a set of primer/probe was developed for detection of PCV 2. In this study we applied real-time PCR assay to 320 samples, collected from pig farms. In 151 of 320 samples, PCV 2 DNA was detected by conventional PCR assay. All samples positive for PCV 2 DNA in conventional PCR assay were also positive in Real-time PCR assay, but 69 of 169 samples that tested negative for PCV 2 DNA in conventional assay were tested positive in TaqMan probe real-time PCR assay. The test of TaqMan probe real-time PCR resulted in detection and quantification limits of 101 copies per sample. TaqMan probe real-time PCR assay increased the number of samples in which PCV 2 was detected by 21%. TaqMan probe real-time PCR assay is very efficient method in contrast to the conventinal PCR, becoming increasingly important method for gene analysis.

Detection of Irradiated Fruits Using the DNA Comet Assay (DNA Comet Assay를 이용한 과일의 방사선 조사 확인)

  • Oh, Kyong-Nam;Park, Jun-Young;Kim, Kyeung-Eun;Yang, Jae-Seung
    • Korean Journal of Food Science and Technology
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    • v.32 no.3
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    • pp.531-537
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    • 2000
  • The simple microgel electrophoresis of single cells, a 'comet assay', on fruit seeds enabled the rapid identification of irradiated fruits by comparing the intact non-irradiated cells and the damaged cells of irradiated fruits. Grapes and plums were irradiated with 0.1, 0.5, 0.7, 1.0 kGy and strawberries, peaches, apples, and nectarines were irradiated with only 1.0 kGy. Seeds were isolated, crushed, and the suspended cells were embedded in an agarose layer. After lysis of the cells, they were subjected to microgel electrophoresis for 2 minutes, and then stained. The DNA radiation-induced fragmentation of all the fruits stretched and migrated out of the cells forming a tail toward the anode giving the appearance of a comet, while the undamaged cells appeared as intact nuclei without tails. Grape and plum seeds irradiated at 0.5 kGy and higher showed significant increases in tail length. With increasing the irradiation doses, longer extention of the DNA from the nucleus toward the anode was observed. Strawberry, peach, apple, and nectarine seeds irradiated with 1.0 kGy also showed the longer tails than non-irradiated ones. DNA comet assay as a rapid and inexpensive screening technique could be an officially validated method for the detection of irradiated fruits.

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