engleski

Evaluation of the heavy metals-induced genotoxicity in plant cells using Comet assay

The Comet assay is a rapid, sensitive and inexpensive method for measuring DNA strand breaks. Relative to other genotoxicity tests, the advantages of the Comet assay include its demonstrated sensitivity for detecting low levels of DNA damage, the requirement for small number of cells per sample and no need for cell division.The ability of Allium cepa L. to accumulate and tolerate cadmium chloride (Cd2+) and thallium acetate (Tl+) at low-doses (1-50 μ M) was tested under continuous growth of several successive harvests of shoots. The potential for DNA strand breaks to act as a biomarker of genetic toxicity in plants in vivo was considered, in view of the fact that higher plants represent reliable sensors in an ecosystem. The nuclear suspension was used in the alkaline Comet assay (pH >13) in order to get minimum DNA migration in the control group and, at the same time, maximum sensitivity for the treated sample. It is concluded that DNA-unwinding time is a critical parameter of the Comet assay and that it may require optimisation for each tissue cell. An extended period of cell lysis up to 24 h failed to produce comet tails in control cells. This indicates that the effects reported here are associated with exposure to the strongly alkaline conditions used in the DNA-unwinding stage of the assay. Comets with different shapes reflecting stages of DNA fragmentation in plant cell of A. cepa exposed to heavy metals indicate Cd2+ less genotoxic than Tl+.

Comet assay; thallium acetate; Allium cepa

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