Logic Circuits for Intelligent Microcystin Monitoring Based on Aptamer Recognition and Toehold-Mediated Hairpin DNA Self-Assembly

A sensitive fluorescence biosensor was developed for microcystin-LR (MC-LR) detection using H1, H2, and H3 DNA probes as sensing elements. The aptamer in H1 can recognize the target. H2 was labeled with FAM and BHQ. The MC-LR and H1 binding will activate the H2 and H3 self-assemblies through toehold-mediated strand displacement. In the formed products (MC-LR/H1/nH2/nH3), FAM and BHQ will be separated and a high FAM fluorescence signal can be observed for the MC-LR assay. The biosensor is sensitive with a detection limit of 53 fM. We further constructed several logic circuits (AND-AND cascaded circuit, feedforward circuit, and resource allocation circuit) using MC-LR, MC-LA, and MC-YR as the three inputs. The numbers 0 and 1 are used to code the input and output signals. The AND-AND cascade circuit can produce a high output signal only in the (111) input combination. In the feedforward circuit, MC-LR and MC-LA can activate the logic circuit to produce high signals, and MC-YR will inhibit the self-assembly and execute the negative feedforward operation. Through the rational design of the DNA probe hybridizations on four different magnetic beads (MBs), the resource allocation circuit can achieve an intelligent allocation of input information. Our proposed fluorescence biosensor can not only provide a sensitive platform for microcystin detection but also serve as a smart and intelligent logic system for microcystin sensing.