The comet assay has proved itself to be not only a method of detection of DNA damages at the level of individual cells but also an approach for investigation of spatial organization of DNA loop domains in nucleoids. Usually, those nucleoids are obtained after cell lysis in high-salt buffer (e. g. 2.5 M NaCl) with a detergent: these conditions ensure the removal of cell membranes and most of the chromatin proteins, while supercoiled DNA loop domains remain untouched. In this work, we tested the comet assay applied to nucleoids obtained in low-salt solution (1 M NaCl). These nucleoids keep most of the histones and thus contain the loops resembling the chromatin loops to a greater extent. It was shown that, despite some quantitative differences, the most general features of the kinetics of DNA exit are about the same for nucleoids obtained in high- and low-salt conditions. It can be concluded that the DNA loops in high-salt nucleoids can be efficiently used to investigate the spatial DNA organization in chromatin.
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Author Name: K. S. Afanasieva, V. V. Olefirenko, A. V. Sivolob
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Keywords: cell lysis, comet assay, DNA loop domains, nucleoids
ISSN: 2409-4943
EISSN: 2413-5003
EOI/DOI: https://doi.org/10.15407/ubj90
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