With　the　biological　significance　and　important　advances　of　nano-scale　DNA　devices,　scientific　activities　have　been　directed　toward　developing　molecular　machinery.　In　this　work,　we　present　a　novel　two-wheel　drive-based　DNA　nanomachine　composed　of　one　signaling　recognition　probe　(SRP),　one　label-free　recognition　probe　(LRP),　and　one　driving　primer　(DP).　Target　DNA　hybridization　can　activate　LRP-based　wheel　driving　by　resorting　to　DP-mediated　polymerization/nicking/displacement　cycles.　This　in　turn　results　in　the　accumulation　of　nicked　strand　1　(NS1)　that　can　initiate　extended　SRP-based　wheel　driving.　As　a　result,　the　hairpin　structure　of　SRP　is　stretched　and　pre-quenched　fluorescence　is　restored.　Meanwhile,　lots　of　nicked　strand　2　(NS2)　are　produced,　which　could　hybridize　perfectly　with　SRP　and　lead　to　further　fluorescence　amplification.　It　is　worth　noting　that,　because　the　nanomachine　operation　relies　strongly　on　inputted　target　trigger,　the　unwanted　background　is　completely　eliminated.　The　detection　limit　of　1　pM　and　an　excellent　capability　to　recognize　the　single-base　mutation　were　achieved.　Significantly,　the　interrogating　of　target　trigger　extracted　from　cancer　cells　is　already　available,　reflecting　the　potential　for　practical　applications.　As　a　proof-of-concept　building,　the　unique　analytical　properties　would　significantly　benefit　the　DNA　nanomachines　and　reveal　great　promise　in　biochemical　and　biomedical　studies.　(C)　2016　Elsevier　Ltd.　All　rights　reserved.