A high sensitivity strategy of DNMT1 activity detection based on self-assembled nucleic acid probe signal amplification technique

Abstract

The abnormal expression of DNA methyltransferase1 (DNMT1) leads to change of genome methylation pattern, which may result in the occurrence and development of cancer. Therefore, the determination of DNMT activity is of great significance for cancer diagnosis and drug screening. In this study, we developed a novel method to detect DNMT1 activity using self-assembly nucleic acid probe signal amplification based on a fluorescence assay. The semi-methylated Biotin-S1'-S2' with sticky ends was fixed to the magnetic beads through the affinity of streptavidin and Biotin. Afterward, pre-prepared poly Tetramethylrhodamine (TAMRA) was added for base complementary pairing with Biotin-S1'-S2' via its sticky end when DNMT1 is present and BssHII shearing is blocked by fully methylated bilayers, thus, the amplified fluorescent signal can be detected. The results showed that the fluorescence intensity of the system was positively correlated with the concentration of DNMT1 in the concentration range of 1–100 nmol/L, and the detection limit was as low as 0.5 nmol/L. The method is simple, highly visualized and successfully applied for the recovery of DNMT1 activity in serum samples. Thus, the method shows great potential for application in clinical diagnosis related to DNMT1.