Use of the random amplif|ed polymorphic DNA (RAPD) assay for the detection of DNA damage and mutations: possible implications of confounding factors

Abstract

The aim of this study was to evaluate the potential of the random amplified polymorphic DNA (RAPD) assay to qualitatively detect the kinetics of benzo[a]pyrene (B[Ma]P)-induced DNA effects in the water flea Daphnia magna exposed to 25 and 50 μg l^-1 B[a]P for 7 and 6 days, respectively. Mortality was recorded on a daily basis in both experiments, and RAPD analysis was performed on samples collected every day following isolation of genomic DNA. The main changes occurring in RAPD profiles produced by the population of Daphnia magna exposed to 25 and 50μg l^-1 B[a]P was a decrease and increase in band intensity, respectively. Most of the changes occurring in the RAPD patterns were likely to be the result of B[a]P-induced DNA damage (B[a]P DNA adducts, oxidized bases, DNA breakages) and/or mutations (point mutations and large rearrangements). In addition, reproducible changes also occurred in the profiles generated by control Daphnia magna. The results lead us to suggest that, in addition to B[a]P-induced DNA damage and mutations, factors such as variation in gene expression, steady levels of genetic alterations and changes in metabolic processes could induce some changes in RAPD patterns. Nevertheless, our data suggest that DNA damage and mutations appear to be the main factors influencing RAPD patterns. This study also emphasizes that unexpected variation in control profiles is not always associated with artefacts.

Keywords::

• Rapd
• Dna Damage
• Mutation
• Confounding Factors
• B[A]P
• Daphnia Magna