Ap-seq: A method to measure apurinic sites and small base adducts genome-wide

Abstract

DNA is constantly challenged by chemical modification and spontaneous loss of its bases, which results in apurinic sites (AP-sites). In addition to the direct route, modified bases may be converted into AP-sites through enzymatic removal of the base as part of the base excision repair pathway. Here we present the method AP-seq, which allows enriching and sequencing AP-sites genome-wide. Quantification of DNA recovery (AP-quant) allows for relative quantification of global AP-sites, and AP-site pulldown followed by qPCR (AP-qPCR) allows for site-specific damage assessment. Taking advantage of glycosylases that specifically excise modified bases also in vitro, this method allows not only to address the genomic distribution of AP-sites but also to detect base modifications, e.g., 8-oxo-7,8-dihydroguanine (8-oxoG). AP-quant, AP-qPCR, and AP-seq can be applied to investigate quantitatively the relative amount and genome specificity of DNA damage and repair, effects of radiation, as well as multiple other questions around AP-sites and base modifications.