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首页> 外文期刊>Nucleic acids research >A panel of colorimetric assays to measure enzymatic activity in the base excision DNA repair pathway
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A panel of colorimetric assays to measure enzymatic activity in the base excision DNA repair pathway

机译：一系列比色测定法，用于测量碱基切除DNA修复途径中的酶活性

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DNA repair is essential for the maintenance of genomic integrity, and evidence suggest that inter-individual variation in DNA repair efficiency may contribute to disease risk. However, robust assays suitable for quantitative determination of DNA repair capacity in large cohort and clinical trials are needed to evaluate these apparent associations fully. We describe here a set of microplate-based oligonucleotide assays for high-throughput, non-radioactive and quantitative determination of repair enzyme activity at individual steps and over multiple steps of the DNA base excision repair pathway. The assays are highly sensitive: using HepG2 nuclear extract, enzyme activities were quantifiable at concentrations of 0.0002 to 0.181 μg per reaction, depending on the enzyme being measured. Assay coefficients of variation are comparable with other microplate-based assays. The assay format requires no specialist equipment and has the potential to be extended for analysis of a wide range of DNA repair enzyme activities. As such, these assays hold considerable promise for gaining new mechanistic insights into how DNA repair is related to individual genetics, disease status or progression and other environmental factors and investigating whether DNA repair activities can be used a biomarker of disease risk.
机译：DNA修复对于维持基因组完整性至关重要，证据表明，DNA修复效率的个体差异可能会导致疾病风险。但是，需要一个鲁棒的测定方法来定量测定大型队列和临床试验中的DNA修复能力，以全面评估这些明显的关联。我们在这里描述了一套基于微孔板的寡核苷酸测定方法，用于在DNA碱基切除修复途径的各个步骤和多个步骤中对修复酶活性进行高通量，非放射性和定量测定。该测定法高度敏感：使用HepG2核提取物，每个反应的酶活度可在0.0002至0.181μg的浓度下定量，具体取决于所测量的酶。测定的变异系数可与其他基于微孔板的测定相比。该测定形式不需要专业设备，并且具有扩展用于分析多种DNA修复酶活性的潜力。因此，这些测定法对于获得有关DNA修复与个体遗传，疾病状况或进展以及其他环境因素的关系的新的机械见解以及调查DNA修复活动是否可以用作疾病风险的生物标志物具有广阔的前景。

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