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Genomic oxidation and alkylation are two of the most important forms of cytotoxic damage that may induce mutagenesis, carcinogenicity, and teratogenicity. Human 8-oxoguanine (hOGG1) and alkyladenine DNA glycosylases (hAAG) are responsible for two major forms of oxidative and alkylative damage repair, and their aberrant activities may cause repair deficiencies that are associated with a variety of human diseases, including cancers. Due to their complicated catalytic pathways and hydrolysis mechanisms, simultaneous and accurate detection of multiple repair glycosylases has remained a great challenge. Herein, by taking advantage of unique features of T7-based transcription and the intrinsic superiorities of single-molecule imaging techniques, we demonstrate for the first time the development of a controlled T7 transcription-driven symmetric amplification cascade machinery for single-molecule detection of …