Abstract
We developed a second-generation DNA-based molecular automaton (MAYA-II) consisting of 129 deoxyribozyme-based logic gates distributed across nine wells of a micro-titre plate. MAYA-II plays a complete game of tic-tac-toe against a human opponent according to a perfect (never losing) strategy. The automaton responds to sequential additions of 32 oligonucleotides keyed to all legal human moves, and displays both human and automaton responses using two-colour fluorogenic outputs. The integration of over one hundred molecular logic gates in a single system represents, in an analogy to silicon based systems, the first "medium-scale integrated molecular circuit". MAYA-II belongs to a larger class of oligonucleotideanalyzing automata that can be programmed to distinguish large numbers of oligonucleotides in mixtures by assigning specific binary encoded outputs in real-time. These systems may be useful in the development of rapid diagnostic arrays, such as multiplex SNP detection and viral lineage attribution.