| 主权项 |
1. A method for quantitative assessment of the base excision repair (BER) and nucleotide excision repair (NER) DNA repair capacities of at least one cellular extract, which method comprises the following steps of:
a) preparing a range of plasmids, each comprising distinct DNA lesions, by independent treatment of said 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 said range of plasmids, c) depositing the plasmids of said 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, x corresponding to an integer equal to the number of cellular extracts to be simultaneously tested, each zone A1 to Ax comprising, the following deposits of plasmids: at least one deposit of control plasmid, a deposit of plasmid containing photoproducts, a deposit of plasmid containing oxidative damage, and a deposit of plasmid containing etheno-bases, d) incubating said functionalized support obtained in step (c) with various repair solutions, each of which comprises at least one cellular extract from a subject, labeled nucleotide triphosphates, and a single repair buffer; e) washing said functionalized support at least once, f) directly or indirectly measuring the signal produced by said labeled nucleotide triphosphate incorporated into the DNA during the repair reaction in step (d), in each of said different and pre-established zones A1 to Ax, g) recording and quantifying the signal corresponding to each deposit of plasmid in each zone A1 to Ax, and h) determining the ratio of the signals of the plasmids comprising the lesions relative to the control plasmid jointly deposited. |
