Ultrasensitive label-free amplified colorimetric detection of p53 based on G-quadruplex MBzymes

Abstract

A novel label-free DNAzyme molecular beacon (MBzyme) strategy was for the first time developed for colorimetric amplification detection of target nucleic acids. The MBzyme, which is designed to contain peroxidase-mimicking DNAzyme that is locked by a common hairpin, was engineered to form a catalytically active MBzyme through hybridizing with the target p53 DNA. The MBzyme is a multifunctional label-free probe that can act as the target recognition element, catalytic DNAzyme and the primer of polymerization. The target p53 DNA hybridization can induce the isothermal circular strand-displacement polymerization even without any chemical modification and other DNA sequences. This unique amplifying strategy leads to the generation of multiple numbers of active MBzyme molecules even if one hybridization event occurs, achieving a dynamic range of seven orders of magnitude and giving a detection limit down to 25 fM which is 3-5 orders of magnitude lower than those of related literature reports. These achievements might be helpful in the design of highly efficient enhancers for G-quadruplex-hemin DNAzymes to be applied on the fundamental research, biotechnology, and biomedical diagnosis.

Keywords: G-quadruplex; Hemin; Isothermal circular strand-displacement polymerization; MBzyme; Ultrasensitive colorimetric assay; p53 gene.