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.

C9orf72 nucleotide repeat structures initiate molecular cascades of disease

“…A colorimetric assay was performed to identify the formation of RNA G-quadruplexes utilizing the enzyme-like peroxidase activity of G-quadruplex•hemin complexes 29 . b) Workflow considerations for the transcriptional assay.…”

Double-stem Hairpin Probe and Ultrasensitive Colorimetric Detection of Cancer-related Nucleic Acids

“…It is clear and unambiguous that the newly-proposed DHP colorimetric assay can offer a very exciting LOD at least 20 times lower than the literature values. 29 , 37 - 43 The measured data demonstrates that this homogenous signaling platform is efficient for ultrasensitive colorimetric detection of DNA hybridization.…”

“…It is worth pointing out that, compared with these sensing systems, 29 , 37 - 43 besides desirable LOD, the dynamic response range is substantially widened, including two linear calibration curves different from each other in the low and high target concentration. Figure 3 C displays a good linear relationship between the peak absorption value (Y) and the logarithmic concentration of target DNA (X) in the range from 1.0 fM to 1 nM with a calibration curve fitted to the equation: Y= 0.340+ 0.0049 LogX (R 2 = 0.9910).…”

Dual‐Hemin‐Labelled Catalytic Molecular Beacon: A Monomer‐Dimer Switching Probe for Sensitive Chemiluminescence Detection of Biomolecules

“…Due to produce the multiple signal molecules through an enzymatic cycle, the DNAzymes‐based MBs exhibit much higher sensitivity than the traditional MB . As an important DNAzymes, G‐quadruplex‐hemin DNAzyme can H 2 O 2 ‐mediated oxidation reaction like peroxidase, and has been widely combined with MBs to develop a catalytic DNA probe for DNA and other biomolecule detection ,. Although G‐quadruplex‐based DNAzyme has been well studied, the preparation of outstanding peroxidase‐like DNAzyme remains challenging.…”

“…Although G‐quadruplex‐based DNAzyme has been well studied, the preparation of outstanding peroxidase‐like DNAzyme remains challenging. First, the bind of G‐quadruplex with hemin is rather slow, and needs 40 min to 1 h ,. So, the entire procedure for preparation of G‐quadruplex‐based DNAzyme is relatively time‐consuming.…”