Application of the comet assay to measure DNA damage induced by UV radiation in the hydrophyte, Spirodela polyrhiza

Abstract

The single-cell gel electrophoresis or comet assay is now widely used to detect DNA damage in animal cells induced by radiation or chemicals. Here, we apply the comet assay to measure ultraviolet (UV)-B-induced DNA damage in plant cells. The accepted animal cell protocol for the comet assay was modified to adapt it to plant cells. The major modifications were conversion of the plant cells to protoplasts and the use of T4 endonuclease V. As a positive control hydrogen peroxide was applied. Significant DNA damage was detected at 100 μM H2O2. This type of DNA damage was not affected by T4 endonuclease V treatment, which implies that the mechanism of H 2O2-induced DNA damage was different from UV-B-induced DNA damage. Our results also indicate that both UV-A and UV-B radiation can induce DNA single-strand breaks in plant cells, while UV-B was more effective than UV-A for inducing pyrimidine dimer formation. © Physiologia Plantarum 2007.