Evaluation of UV damage at DNA level in <Emphasis Type="Italic">Nicotiana plumbaginifolia</Emphasis> protoplasts using single cell gel electrophoresis |
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Authors: | Yulheri Abas Nadia Touil Micheline Kirsch-Volders Geert Angenon Michel Jacobs Ivan Diane Hector Famelaer |
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Institution: | (1) Laboratory of Plant Genetics, Department of Applied Biological Sciences, Institute of Molecular Biology and Biotechnology, Vrije Universiteit Brussel, Pleinlaan 2, 1050 Brussels, Belgium;(2) Centre for Forest Plantation Research and Development, Department of Forestry, Jl. Monjali, Pakem, Sleman, Yogyakarta, Indonesia;(3) Laboratory of Cell Genetics, Department of Biological Sciences, Vrije Universiteit Brussel, Pleinlaan 2, 1050 Brussels, Belgium |
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Abstract: | The aim of the present study was to observe the induction and repair of single strand breaks (Ssbs) and double strand breaks
(Dsbs) in mesophyll protoplasts of Nicotiana plumbaginifolia, irradiated with UV-C and cultured under light or dark conditions. DNA damage and repair was determined by the neutral and
alkaline comet assay to reveal Dsbs and Ssbs respectively.
Subculturing protoplasts for 4 h at low temperature was essential to reduce the amount of Dsbs to the detection limit of the
assay procedure.
Light-cultured protoplasts showed a significant increase of Ssbs and Dsbs compared to dark cultured protoplasts, in which
the number of Ssbs and Dsbs remained very constant throughout the experiments.
UV treatment significantly enhanced the levels of Ssbs and Dsbs in light and dark cultured protoplasts. On average, equal
levels of DNA damage were observed under light or dark conditions. Formulations introduced to evaluate the contribution of
UV-C or light treatment in repair kinetics of DNA damage, showed that the number of Ssbs, but not of Dsbs, evolved differently
for light and dark cultured protoplasts. DNA repair was more rapidly observed under light conditions and occurred in different
repair phases. Observations are discussed in relation to the involvement of chromatin remodelling, photosynthetic active radiation
and DNA repair mechanisms. |
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Keywords: | Comet assay DNA damage and repair Plant protoplast UV-C irradiation |
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