Optimization of the 32 P-postlabeling Assay for Detecting Benzo(a)pyrene-induced DNA Adduct Formation in Zacco platypus

• 영문명: Optimization of the 32 P-postlabeling Assay for Detecting Benzo(a)pyrene-induced DNA Adduct Formation in Zacco platypus
• 발행기관: 한국환경보건학회
• 저자명: Jin Wuk Lee Sung Kyu Lee
• 간행물 정보: 『한국환경보건학회지』제40권 제1호, 55~62쪽, 전체 8쪽
• 주제분류: 공학 > 환경공학
• 파일형태: PDF
• 발행일자: 2014.02.28

영문 초록

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.

목차

I. Introduction

II. Materials and Methods

III. Results

IV. Discussion

V. Conclusions

키워드

P-postlabeling assay Benzo(a)pyrene Zacco platypus