An integrated and restructive probe mediated strand displacement amplification strategy for sensitive and specific DNA methyltransferase activity detection

Herein, an integrated and restructive probe mediated strand displacement amplification (SDA) strategy is developed for sensitive and specific DNA MTase activity detection. The probe is specially designed with a hairpin structure, which encloses the MTase recognition site in the stem and seals the SDA primer and template in the loop. Under the action of DNA MTase, the probe is methylated and cleaved by DpnI endonuclease, causing its stem truncated. The truncated structure then reconstructs with a shrunken loop and a partially-hybridized duplex of stem, drawing the SDA primer and template domains close to trigger cascade amplification. Numerous G-quadruplexes are produced and intercalated by N-methyl-mesoporphyrin IX (NMM) to generate enhanced fluorescent signal. This strategy detects MTase activity down to 0.063 U/mL, and well distinguishes Dam MTase from its analogues. It is further applied for the MTase detection in biological samples and MTase inhibition analysis. The strategy will provide a promising tool for detection MTase activity in biomedical study and early cancer diagnosis. (C) 2018 Elsevier B.V. All rights reserved.