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http://hdl.handle.net/1942/33016
Title: | An optimized comet-based in vitro DNA repair assay to assess base and nucleotide excision repair activity | Authors: | Vodenkova, Sona Azqueta, Amaya Collins, Andrew Dusinska, Maria Gaivao, Isabel Moller, Peter Opattova, Alena Vodicka, Pavel Godschalk, Roger W. L. LANGIE, Sabine |
Issue Date: | 2020 | Publisher: | NATURE RESEARCH | Source: | NATURE PROTOCOLS, 15 (12) , p. 3844 -3878 | Abstract: | This optimized protocol (including links to instruction videos) describes a comet-based in vitro DNA repair assay that is relatively simple, versatile, and inexpensive, enabling the detection of base and nucleotide excision repair activity. Protein extracts from samples are incubated with agarose-embedded substrate nucleoids ('naked' supercoiled DNA) containing specifically induced DNA lesions (e.g., resulting from oxidation, UVC radiation or benzo[a]pyrene-diol epoxide treatment). DNA incisions produced during the incubation reaction are quantified as strand breaks after electrophoresis, reflecting the extract's incision activity. The method has been applied in cell culture model systems, human biomonitoring and clinical investigations, and animal studies, using isolated blood cells and various solid tissues. Once extracts and substrates are prepared, the assay can be completed within 2 d. This protocol describes a comet-based in vitro assay for detecting base and nucleotide excision repair activity for use in cell culture model systems, human biomonitoring and clinical investigations, and animal studies, using isolated blood cells and various solid tissues. | Notes: | Langie, SAS (corresponding author), Maastricht Univ, Sch Nutr & Translat Res Metab NUTRIM, Dept Pharmacol & Toxicol, Maastricht, Netherlands.; Langie, SAS (corresponding author), Hasselt Univ, Ctr Environm Sci, Hasselt, Belgium. s.langie@maastrichtuniversity.nl |
Other: | Langie, SAS (corresponding author), Maastricht Univ, Sch Nutr & Translat Res Metab NUTRIM, Dept Pharmacol & Toxicol, Maastricht, Netherlands ; Hasselt Univ, Ctr Environm Sci, Hasselt, Belgium. s.langie@maastrichtuniversity.nl | Keywords: | Oxidatively Damaged Dna;Cancer Patients;Polyphenolic Compounds;Occupational-Exposure;Incision Activity;Expression Levels;Telomere Length;Mineral Fibers;Cross-Links;Cells | Document URI: | http://hdl.handle.net/1942/33016 | ISSN: | 1754-2189 | e-ISSN: | 1750-2799 | DOI: | 10.1038/s41596-020-0401-x | ISI #: | WOS:000590016600002 | Rights: | 2020 Springer Nature Limited. | Category: | A1 | Type: | Journal Contribution | Validations: | ecoom 2021 |
Appears in Collections: | Research publications |
Files in This Item:
File | Description | Size | Format | |
---|---|---|---|---|
Vodenkova+et+al_Nature+Protocols_2020_Postprint.pdf | Peer-reviewed author version | 2.59 MB | Adobe PDF | View/Open |
s41596-020-0401-x.pdf Restricted Access | Published version | 3.43 MB | Adobe PDF | View/Open Request a copy |
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