METHOD FOR THE QUANTITATIVE ASSESSMENT OF GLOBAL AND SPECIFIC DNA REPAIR CAPACITIES OF AT LEAST ONE BIOLOGICAL MEDIUM, AND THE APPLICATIONS THEREFOR

摘要

A method for quantitative assessment of base excision repair (BER) and nucleotide excision repair (NER) DNA repair capacities of at least one cellular extract, which method entails: a) preparing a range of plasmids with at least one physical or chemical treatment or both and recovering a supercoiled fraction of each of said plasmids, b) characterizing the lesions present on each of the plasmids of the range of plasmids; c) depositing the plasmids of the range of plasmids, and at least one supercoiled control plasmid without lesions onto a single solid support, according to a pre-established configuration A, so as to form a functionalized support divided into different zones A1 to Ax, corresponding to an integer equal to the number of cellular extracts to be simultaneously tested, each zone containing supercoiled plasmids treated with at least one physical or chemical treatment that induces a lesion acted upon by BER mechanisms, and at least one other zone containing supercoiled plasmids treated with at least one physical or chemical treatment that induces a lesion acted upon by NER mechanisms; d) incubating the functionalized support obtained in step c) with various repair solutions, each of which contains at least one cellular extract from a subject, and labeled nucleotide phosphates; e) washing the functionalized support at least once; f) directly or indirectly measuring the signal produced by the labeled nucleotide triphosphate incorporated into the DNA during the repair reaction in step d), in each of the different and pre-established zones A₁ to A_x; g) recording and quantifying the signal corresponding to each deposit of plasmid in each zone A₁ to A_x; and h) determining the ratio of the signals of the plasmids containing the lesions relative to the control plasmid jointly deposited.