DNA methyltransferase activity detection based on fluorescent silver nanocluster hairpin-shaped DNA probe with 5’-C-rich/G-rich-3’ tails

Abstract DNA methylation has received a large amount of attention due to its close relationship to a wide range of biological phenomena, such as gene activation, gene imprinting, and chromatin stability. Herein, we have designed a hairpin-shaped DNA probe with 5’-C-rich/G-rich-3’ tails and developed a simple and reliable fluorescence turn-off assay for DNA adenine methylation (Dam) methyltransferase (MTase) detection combining site recognition and the fluorescence enhancement of DNA-templated silver nanoclusters (DNA–AgNCs) by guanine-rich DNA sequences. A designed hairpin probe with 5’ CCCTTACCCC and 3’ GGGTGGGGTGGGGTGGGG displays a bright red emission after reacting with AgNO 3 and NaBH 4 . In the presence of Dam MTase, the methylation-sensitive restriction endonuclease Dpn I which has the same recognition site with the Dam MTase can split the probe, freeing the G-rich sequence from the C-rich sequence, thus quenching the fluorescence of DNA–AgNCs. Compared to traditional fluorescent-based methods, this strategy is simple and inexpensive. A linear response to concentrations of Dam MTase which range from 1 U/mL to 100 U/mL and a detection limit of 1 U/mL are obtained without any amplification steps. In addition, we also demonstrate the method can be used for evaluation and screening of inhibitors for Dam MTase.