Cascaded molecular logic gates using antibiotics as inputs based on exonuclease III and DNAzyme

We successfully constructed several cascaded molecular logic gates (2INHIBIT-2AND, 2AND-2OR, and 2OR-2INHIBIT) using three different antibiotics as the inputs. In the presence of kanamycin (KAN), chloramphenicol (CHL), or oxytetracycline (OXY), the aptamer-antibiotic recognition will release the trigger DNA to active the hairpin DNA hybridization. Exonuclease III (Exo III)-mediated catalysis reaction was introduced in the logic system to generate Mg²⁺-dependent DNAzyme, which was used to cleave the fluorescence signal reporter probe. For input, the presence and absence of the antibiotic was defined as 1 and 0, respectively. For output, the fluorescence intensity higher or lower than the threshold value was defined as 1 and 0, respectively. In the 2INHIBIT-2AND logic circuit, the 101 input combination generates an output of 1 and other input combinations generate an output of 0. In the 2AND-2OR logic circuit, the input combinations of 001, 011, 110, 101, and 111 generate an output of 1 and other input combinations generate an output of 0. In the 2OR-2INHIBIT logic circuit, the input combinations of 010, 100, and 110 generate an output of 1 and other input combinations generate an output of 0. Our constructed logic system exhibits high selectivity and can work even in complex water samples. With the advantages of multiple biocomputation capabilities, high flexibility, and easy scalability, this logic gate system provides a new analytical method for the intelligent detection of different antibiotics.