DOI QR코드

DOI QR Code

Application of Single Cell Gel Electrophoresis for Detection of DNA Single Strand Breaks in DNA of Fish Blood Cell

어류혈구세포에 있어서 Single Cell Gel Electrophoresis를 응용한 DNA Single Strand Breack의 측정

  • KIM Gi Beum (Department of Marine Environmental Engineering / The Institute of Marine Industry, Gyeongsang National University) ;
  • LEE Richard F. (Skidaway Institute of Oceanography) ;
  • MARUYA Keith A. (Skidaway Institute of Oceanography)
  • 김기범 (경상대학교 해양환경공학과 / 해양산업연구소) ;
  • ;
  • Published : 2003.08.01

Abstract

Single-cell gel electrophoresis (comet assay) was used to detect DNA single strand break in blood cells from several marine fish species. Three fish species were collected from Georgia coastal area. Mummichog, Fundulus heteroclitus showed higher DNA damage than sea bass, Lateolabrax japonicus and trout, Oncorhynchus masou masou under the same experimental conditions. Mummichogs had more alkaline-labile sites on their DNA than other fish species. The comet assay with mummichog blood cells at pH 12.5 showed a dose-response curve with the increasing concentrations of hydrogen peroxide. While the isolated leucocytes showed no increase of DNA damage after in vitro exposure to 2-methyl-1,4-naphthoquinone (MNQ), erythrocytes showed dose-dependent DNA damage. These results indicate that the comet assay can be applied successfully as a bioassay using erythrocyte for environmental monitoring.

Keywords

References

  1. $Ban\dot ath$, J.P., M. Fushiki and P.L. Olive. 1998. Rejoining of DNA single- and double-strand breaks in human white blood cells exposed to ionizing radiation. Int. J. Radiat. BioI., 73, 649-660 https://doi.org/10.1080/095530098141906
  2. Gedik, C.M., S.W.B. Ewen and A.R. Collins. 1992. Single-cell gel electrophoresis applied to the analysis of UV-C damage and its repair in human cells. Int. J. Radiat. BioI. 62, 313-320 https://doi.org/10.1080/09553009214552161
  3. $Horv\dot athov\dot a$, E., D. $Slamenov\dot a$, L. Hlincikova, T.K. Mandal, A. $G\dot abelov\dot a$ and A.R. Collins. 1998. The nature and origin of DNA single-strand breaks determined with the comet assay. Mutation Res., 409, 163-171 https://doi.org/10.1016/S0921-8777(98)00053-6
  4. Kammann, U., J.C. Riggers, N. Theobald and H. Steinhart. 2000. Genotoxic potential of marine sediments from the North Sea. Mutation Res., 467, 161-168 https://doi.org/10.1016/S1383-5718(00)00030-9
  5. Kasamatsu, T., K. Kohda and Y. Kawazoe. 1996. Com-parison of chemically induced DNA breakage in cellular and subcellular systems using the comet assay. Mutation Res., 369, 1-6 https://doi.org/10.1016/S0165-1218(96)90041-5
  6. Kim, G.B. and R.F. Lee. Effects of genotoxic compounds on DNA and development of early and late grass shrimp embryo stages. Mar. Environ. Res. (submitted)
  7. Kim, G.B., R.F. Lee and D.L. Mitchell. 2000. Damage to grass shrimp (Palaemonetes pugio) embryo DNA by summer sunlight followed by DNA repair in the dark. Mar. BioI., 137, 675-682 https://doi.org/10.1007/s002270000371
  8. Koppen, G. and K.J. Angelis. 1998. Repair ofX-ray induced DNA damage measured by the comet assay in roots of Vicia faba. Environ. Mol. Mutagenesis, 32, 281-285 https://doi.org/10.1002/(SICI)1098-2280(1998)32:3<281::AID-EM11>3.0.CO;2-R
  9. Kurelec, B. 1993. The genotoxic disease syndrome. Mar. Environ. Res., 35, 341-348 https://doi.org/10.1016/0141-1136(93)90101-5
  10. Lee, R.F. and G.B. Kim. 2002. Comet assay to assess DNA damage and repair in grass shrimp embryos exposed to phototoxicants. Mar. Environ. Res., 54,465-469 https://doi.org/10.1016/S0141-1136(02)00128-9
  11. Lee, R.F, G.B. Kim, K.A. Maruya, S.A. Steinert and Y. Oshima. 2000. DNA strand breaks (comet assay) and embryo development effects in grass shrimp (Palae-monetes pugio) embryos after exposure to genoto-xicants. Mar. Environ. Res., 50, 553-557 https://doi.org/10.1016/S0141-1136(00)00110-0
  12. Lee, R.F., S.A. Steinert, K. Nakayama and Y. Oshima. 1999. Use of DNA strand damage (comet assay) and embryo hatching effects to assess contaminant ex-posure in blue crab (Callinectes sapidus) embryos. In: Environmental Toxicology and Risk Assessment, Vol 8. ASTM STP 1364, D. Hensel, M.C. Black, eds. American Society of Testing and Materials, West Conshohocken, PA. pp. 341-349
  13. McKelvey-Martin, V.J., M.H.L. Green, P. Schmezer, B.L. Pool-Zobel, M.P. De Meo and A. Collins. 1993. The single cell gel electrophoresis assay (comet assay): a European review. Mutation Res., 288, 47-63 https://doi.org/10.1016/0027-5107(93)90207-V
  14. Michelmore, C.L., C. Birmelin, D.R. Livingstonr and J.K. Chipman. 1998. Detection of DNA strand breaks in isolated mussel (Mytilus edulis L.) digestive gland cells using the 'comet' assay. Ecotoxicol. Environ. Safety, 41, 51-58 https://doi.org/10.1006/eesa.1998.1666
  15. Mohankumar, M.N., S. Janani, B.K. Prabhu, P.R.V. Kumar and R.K. Jeevanram. 2002. DNA damage and integrity ofUV-induced DNA repair in lymphocytes of smokers analysed by the comet assay. Mutation Res., 520, 179-187 https://doi.org/10.1016/S1383-5718(02)00201-2
  16. Morrison, H., B. Jemstrom, M. Nordenskjold, H. Thor and S. Orrenius. 1984. Induction of DNA damage by menadione (2-methyl-1,4-naphthoquinone) in primary cultures of rat hepatocytes. Biochem. Phar-macol., 33, 1763-1769 https://doi.org/10.1016/0006-2952(84)90347-2
  17. Nacci, D.E., S. Cayula and E. Jackim. 1996. Detection of DNA damage in individual cells from marine organisms using the single cell gel assay. Aquat. Toxicol., 35, 197-210 https://doi.org/10.1016/0166-445X(96)00016-1
  18. Nigro, M., G. Grenzilli, V. Scarcelli V., S. Gorbi and F. Regoli. 2002. Induction of DNA strand breakage and apoptosis in the eel Anguilla anguilla. Mar. Environ. Res., 54, 517-520 https://doi.org/10.1016/S0141-1136(02)00178-2
  19. Pandrangi, R., M. Petras, S. Ralph and M. Vrzoc. 1995. Alkaline single cell gel (comet) assay and genotoxicity monitoring using bullheads and carps. Environ. Mol. Mutagenesis, 26, 345-356 https://doi.org/10.1002/em.2850260411
  20. Shugart, L.R. 1988. Quantitation of chemically induced damage to DNA of aquatic organisms by alkaline unwinding assay. Aquat. Toxicol., 13, 43-52 https://doi.org/10.1016/0166-445X(88)90071-9
  21. Singh, N.P., M.T. McCoy, R.R. Tice and E.L. Schneider. 1988. A simple technique for quantitation oflow levels of DNA damage in individual cells. Exp. Cell Res., 175, 184-191 https://doi.org/10.1016/0014-4827(88)90265-0
  22. Steinert, S.A., R.S. Montee and M.P. Sastre. 1998. Influence of sunlight on DNA damage in mussels exposed to polycyclic aromatic hydrocarbons. Mar. Environ. Res., 46, 355-358 https://doi.org/10.1016/S0141-1136(97)00088-3
  23. Thor, H., M.T. Smith, P. Hartzell, G. Bellomo, S.A. Jewell, S. Orrenius. 1982. The metabolism of menadione (2-methyl-1 ,4-naphthoquinone) by isolated heato-cytes: a study of the implication of oxidative stress in intact-cells. J. BioI. Chem., 257, 2419-2425
  24. $\ddot Undeger$, $\ddot U$. and N. Basaram. 2002. Assessment of DNA damage in workers occupationally exposed to pesti-cide mixtures by the alkaline comet assay. Arch. Toxicol., 76, 430-436 https://doi.org/10.1007/s00204-002-0355-5
  25. Zhu, C.Y. and S. Loft. 2001. Effects of Brussels sprouts extracts on hydrogen peroxide-induced DNA strand breaks in human lymphocytes. Food Chem. Toxicol., 39, 1191-1197. https://doi.org/10.1016/S0278-6915(01)00061-8

Cited by

  1. Genotoxicity (DNA damage) on Blood Cells of Parrot Fish (Oplegnathus fasciatus) Exposed to Acidified Seawater Making of CO2 vol.23, pp.3, 2014, https://doi.org/10.5322/JESI.2014.23.3.483
  2. 유전독성물질로 오염된 해양생물의 생물검정법으로서 comet assay 이용 vol.14, pp.11, 2003, https://doi.org/10.5322/jes.2005.14.11.1071