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Optimization of the 32P-postlabeling Assay for Detecting Benzo(a)pyrene-induced DNA Adduct Formation in Zacco platypus
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  • Optimization of the 32P-postlabeling Assay for Detecting Benzo(a)pyrene-induced DNA Adduct Formation in Zacco platypus
  • Optimization of the 32P-postlabeling Assay for Detecting Benzo(a)pyrene-induced DNA Adduct Formation in Zacco platypus
저자명
Lee. Jin Wuk,Lee. Sung Kyu
간행물명
韓國環境保健學會誌
권/호정보
2014년|40권 1호|pp.55-62 (8 pages)
발행정보
한국환경보건학회
파일정보
정기간행물|ENG|
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이 논문은 한국과학기술정보연구원과 논문 연계를 통해 무료로 제공되는 원문입니다.
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기타언어초록

Objectives: $^{32}P$-postlabeling assay is the most sensitive method of detecting DNA adduct formation. However, it is limited by a low sample throughput and use of radioisotopes (RI). In this study, we modified it to minimize these limitations and applied it to Z. platypus exposed to Benzo(a)pyrene (BaP) in order to investigate DNA adduct formation (effect biomarker for pollutants) in Z. platypus for assessing risk of waterborne BaP exposure. Methods: DNA hydrolysis was performed only with Micrococcal nuclease (MNase), RI reduction test was performed and the overlapping steps between thin layer chromatography (TLC) and radioisotope high-performance liquid chromatography (RI-HPLC) were omitted. The application of a modified method to Z. platypus exposed to BaP was performed. Results: The results revealed that the amount of RIs used can be reduced roughly 10-fold. Because the analysis time was shortened by 8.5 hours, the sample throughput per hour was increased compared with the previous method. The results of applying modified $^{32}P$-postlabeling assay to Z. platypus, DNA adduct formation in Z. platypus showed dose-dependency with the BaP concentration. Only BPDE-dGMP was detected as a DNA adduct. Conclusion: These results demonstrate that the modified $^{32}P$-postlabeling assay is a suitable method for detecting DNA adduct formation in Z. platypus exposed to waterborne BaP and will be useful in risk assessment of carcinogenic effect in aquatic environment due to BaP.