A label-free colorimetric detection of microRNA via G-quadruplex-based signal quenching strategy

Highlights: • A simple and label-free colorimetric biosensing system for the detection of miRNAs. • Utilization of the signal quenching strategy with G-quadruplex DNAzyme. • Considerably low detection limit with naked eyes. • Base-mismatch discrimination among the miRNA family members. -- Abstract: Signal amplification strategy have been the intensive direction for highly-sensitive microRNA (miRNA) detection, however, it is worth noting that signal quenching approach is rarely realized. Inspired by the unwinding of G-quadruplex with Klenow Fragment polymerase (KF), a simple and label-free signal quenching system for detection of miRNAs was developed. In the presence of target miRNA, miRNA was used as a primer to promote KF activity resulting in the disruption of G-quadruplex structure of probe and the biological catalysis inactivation of DNAzyme formed by G-quadruplex structure and hemin. As a result, the probe would be transformed from G-quadruplex to duplex and the obvious differential signal could be observed in most cases even with naked eyes. With the significant signal decreasing, the target miRNA can be rapidly analyzed by UV–vis spectra with a considerably low detection limit (4.5 nM). This method exhibits excellent selectivity and anti-interference ability by discriminating base-mismatched and other miRNA families from target miRNA. Meanwhile, a good recovery (90.7%∼102.4%) in 5% human serum was obtained and this strategy verified a high expression level of miR21 in total RNA of MCF-7 compared with 293T and HeLa. It implies that this assay is of great potential to be applied in biochemical research and clinical diagnosis.