Electrochemical screening of single nucleotide polymorphisms with significantly enhanced discrimination factor by an amplified ratiometric sensor

Abstract

The detection of single nucleotide polymorphisms (SNPs) is of great clinical significance to the diagnosis of various genetic diseases and cancers. In this work, the development of an ultrasensitive ratiometric electrochemical sensor for screening SNP with a significantly enhanced discrimination factor is reported. The ferrocene (Fc) and methylene blue (MB) dual-tagged triple helix complex (THC) probes are self-assembled on the gold electrode to construct the sensing interface. The addition of the mutant p53 gene causes the disassembly of the THC probes with the release of the Fc-tagged sequence and the folding of the MB-labeled sequence into a hairpin structure, causing the change in the current response ratio of MB to Fc for monitoring the mutant p53 gene. Such ratio is dramatically enhanced by the toehold-mediated displacement reaction-assisted target recycling amplification with the presence of an assistance hairpin sequence. With the significant signal amplification and the advantageous specificity of the THC probes, sub-femtomolar detection limit and a highly enhanced SNP discrimination factor for the mutant p53 gene can be obtained. Besides, the proof-of-demonstration application of the sensor for diluted real samples has been verified, offering such sensor new opportunities for monitoring various genetic related diseases.