The development of a versatile biosensing platform to screen specific DNA sequences is still an essential issue of molecular biology research and clinic diagnosis of genetic disease. In this work, we for the first time reported a double-stem hairpin probe (DHP) that was simultaneously engineered to incorporate a DNAzyme, DNAzyme's complementary fragment and nicking enzyme recognition site. The important aspect of this hairpin probe is that, although it is designed to have a long ds DNA fragment, no intermolecular interaction occurs, circumventing the sticky-end pairing-determined disadvantages encountered by classic molecular beacon. For the DHP-based colorimetric sensing system, as a model analyte, cancer-related DNA sequence can trigger a cascade polymerization/nicking cycle on only one oligonucleotide probe. This led to the dramatic accumulation of G-quadruplexes directly responsible for colorimetric signal conversion without any loss. As a result, the target DNA is capable of being detected to 1 fM (six to eight orders of magnitude lower than that of catalytic molecular beacons) and point mutations are distinguished by the naked eye. The described DHP as a-proof-of-concept would not only promote the design of colorimetric biosensors but also open a good way to promote the diagnosis and treatment of genetic diseases.
Keywords: DNAzyme; colorimetric sensing; double-stem hairpin probe (DHP); p53 gene.